Toll-like receptor agonists

ABSTRACT

The present invention relates to imidazo-pyridinyl compounds, or a pharmaceutically acceptable salt thereof, to pharmaceutical compositions comprising such compounds and salts, and to methods of using such compounds, salts and compositions for the treatment of abnormal cell growth, including cancer, in a subject.

BACKGROUND OF THE INVENTION

Toll-like receptors (TLRs) are a family of transmembrane proteins thatrecognize structurally conserved molecules that are derived from andunique to pathogens, referred to as pathogen-associated molecularpatterns (PAMPS). As such, TLRs function in the mammalian immune systemas front-line sensors of pathogen-associated molecular patterns,detecting the presence of invading pathogens (Takeuchi and Akira 2010Cell 140:805-820). TLR engagement in sentinel immune cells causesbiosynthesis of selected cytokines (e.g., type I interferons), inductionof costimulatory molecules, and increased antigen presentation capacity.These are important molecular mechanisms that activate innate andadaptive immune responses. Accordingly, agonists and antagonists of TLRsfind use in modulating immune responses. TLR agonists are typicallyemployed to stimulate immune responses, whereas TLR antagonists aretypically employed to inhibit immune responses (Gosu et al 2012.Molecules 17:13503-13529).

The human genome contains 10 known TLRs, of these TLR3, TLR7, TLR8, andTLR9 recognize nucleic acids and their degradation products. Thedistribution of TLR7, TLR8, and TLR9 is restricted to the endosomalcompartments of cells and they are preferentially expressed in cells ofthe immune system. In the activated dimeric receptor configuration TLR7and TLR8 recognize single strand RNA at one ligand binding site and theribonucleoside degradation products guanosine and uridine, respectively,(as well as small molecule ligands with related structural motifs) at asecond ligand binding site (Zhang et al 2016 Immunity 45(4); 737-748:Tanji et al 2015 Nat Struct Mol Biol 22: 109-115).

Some small-molecule TLR7 or TLR8 agonists have been identified. Thoseagonists can be grouped into purine-like molecules, such as7-thia-8-oxoguanosine (TOG, isatoribine) or the imidazoquinoline-basedcompounds such as imiquimod. Imiquimod is so far the only approved TLR7agonist, marketed as a 5% cream (Aldara). It generates approximately 80%5-year clearance of superficial basal cell carcinomas, which is the mostcommon cancer worldwide, thus demonstrating the importance of TLR7agonists in cancer immunotherapy. The functional expression of TLR7appears to be restricted to specific immune cells. Engagement of TLR7 inplasmacytoid dendritic cells leads to the induction of interferon α/β,which plays essential functions in the control of the adaptive immuneresponse (Bao and Liu 2013 Protein Cell 4:40-5).

Engagement of TLR8 in myeloid dendritic cells, monocytes andmonocyte-derived dendritic cells induces a prominent pro-inflammatorycytokine profile, characterized by increased production of tumornecrosis factor-α, interleukin-12, and IL-18 (Eigenbrod et al J Immunol,2015, 195, 1092-1099).

Small molecule TLR agonists have also been investigated for use asvaccine adjuvants (Dowling, ImmunoHorizons 2018, 2(6) 185-197).

Thus, virtually all major types of monocytic and dendritic cells can beactivated by agonists of TLR7 and TLR8 to become highly effectiveantigen-presenting cells, thereby promoting an effective innate andadaptive immune response. Most antigen presenting cell types expressonly one of these two receptors, accordingly small molecules with potentagonist activity against both TLR7 and TLR8 receptors are potentiallymore effective immune adjuvants than TLR7 agonists alone.

Thus, a TLR7/TLR8 (TLR7/8) small molecule agonist with dual bioactivitycould provide further benefit over a more selective TLR7 agonist andwould cause innate immune responses in a wider range of antigenpresenting cells and other key immune cell types, including plasmacytoidand myeloid dendritic cells, monocytes, and B cells (van Haren et al2016 J Immunol 197:4413-4424; Ganapathi et al 2015 Plos One10(8).e0134640). Such potent dual TLR7/8 agonists may also be effectivein stimulating effective anti-tumor responses in cancer (Singh et al2014 J. Immunol 193 4722-4731: Sabado et al 2015 Cancer Immunol Res 3278-287, Spinetti et al 2016 Oncoimmunol 9; 5(11):e1230578: Patil et al2016 Mini Rev Med Chem 16:309-322).

Despite the success of Imiquimod (Aldera) in treating superficial basalcell carcinoma, there remains a need for not only more potent TLR7agonists, but also balanced, potent TLR7/8 agonists to expand treatmentoptions for patients for various cancers. These treatment options couldbe local administrations which would deliver the drug to the tumordirectly, whilst limiting systemic side effects. Alternatively,systemically administered TLR7 agonists or TLR7/8 agonists would havethe advantage of being able to reach difficult to administer tumors aswell as multiple tumors, through the systemic circulation.

SUMMARY OF THE INVENTION

The present invention provides, in part, compounds of Formula (I),including Formula (Ia) and (Ib), collectively, a compound of theinvention, or a pharmaceutically acceptable salt thereof. Such compoundsactivate the human TLR7 (hTLR7) and also activate the human TLR8(hTLR8), thereby affecting biological functions. In some embodiments,the invention provides compounds that are dual agonists that areselective for both TLR7 and TLR8 (TLR7/8 agonists). In anotherembodiment, the invention provides compounds that are agonists that areselective for TLR7. Another embodiment provides pharmaceuticalcompositions and medicaments comprising the compounds of the invention,or a pharmaceutically acceptable salt thereof, alone or in combinationwith additional anticancer therapeutic agents.

The present invention also provides, in part, methods for preparing thecompounds, pharmaceutically acceptable salts and compositions of theinvention, and methods of using the foregoing alone or in combinationwith additional anticancer therapeutic agents.

In one aspect, the invention provides a compound of Formula (I):

or a pharmaceutically acceptable salt thereof, wherein

R¹ and R² are independently C₁₋₃ alkyl; or

R¹ and R² are joined to form a 5- to 7-membered carbocyclic ring,wherein said carbocyclic ring may be saturated or unsaturated;

R³ is

R⁴ is C₁₋₆ alkyl, or (CH₂)_(n)O(CH₂)_(m)CH₃, wherein the C₁₋₆ alkyl orany carbon of the (CH₂)_(n)O(CH₂)_(m)CH₃ group is substituted with 0 to3 halogen as valency allows;

R⁵ is C₁₋₃ alkyl, or OC₁₋₃ alkyl, wherein the C₁₋₃ alkyl is substitutedby 0 to 3 F;

R⁶ is H, or C₁₋₃ alkyl, wherein the C₁₋₃ alkyl is substituted with 0 to3 F;

m is 0 to 2; and

n is 1 to 3.

In another aspect, the invention provides a pharmaceutical compositioncomprising a compound of the invention, according to any of the formulaedescribed herein, or a pharmaceutically acceptable salt thereof, and apharmaceutically acceptable carrier or excipient. In some embodiments,the pharmaceutical composition comprises two or more pharmaceuticallyacceptable carriers and/or excipients.

In another aspect, the invention also provides therapeutic methods anduses comprising administering a compound of the invention, or apharmaceutically acceptable salt thereof.

In another aspect, the invention provides a method for the treatment ofabnormal cell growth, in particular, cancer, in a subject in needthereof, comprising administering to the subject a therapeuticallyeffective amount of a compound of the invention, or a pharmaceuticallyacceptable salt thereof. Compounds of the invention may be administeredas single agents or may be administered in combination with otheranti-cancer therapeutic agents, including standard of care agentsappropriate for the particular form of cancer. This also includes use ofa compound of the invention, or a pharmaceutically acceptable saltthereof, in the manufacture of a medicament for treating abnormal cellgrowth, in particular, cancer, in a subject in need thereof.

In another aspect, the invention provides a compound of the invention,or a pharmaceutically acceptable salt thereof, for use as a medicament,in particular a medicament for the treatment of abnormal cell growth,such as cancer.

In yet another aspect, the invention provides the use of a compound ofthe invention, or a pharmaceutically acceptable salt thereof, for themanufacture of a medicament for the treatment of abnormal cell growth,such as cancer, in a subject.

In another aspect, the invention includes within its scope thepharmaceutically acceptable salts of the compounds of the invention.Accordingly, the phrase “or a pharmaceutically acceptable salt thereof”is implicit in the description of all compounds described herein unlessexplicitly indicated to the contrary.

DETAILED DESCRIPTION OF THE INVENTION

As provided, the invention concerns a compound of Formula (I) asprovided above.

The present invention may be understood more readily by reference to thefollowing detailed description of additional embodiments of theinvention and the Examples included herein. It is to be understood thatthe terminology used herein is provided for the purpose of describingspecific embodiments only and is not intended to be limiting. It isfurther to be understood that unless specifically defined herein, theterminology used herein is to be given its traditional meaning as knownin the relevant art.

As used herein, the singular form “a”, “an”, and “the” include pluralreferences unless indicated otherwise. For example, “a” substituentincludes one or more substituents. The term “about” means having a valuefalling within an accepted standard of error of the mean, whenconsidered by one of ordinary skill in the art.

The term “alkyl”, as used herein, means a straight or branched chainmonovalent hydrocarbon group of formula —C_(n)H_((2n+1)). Non-limitingexamples include methyl, ethyl, propyl, butyl, 2-methyl-propyl,1,1-dimethylethyl, pentyl and hexyl.

In some instances, the number of carbon atoms in a hydrocarbon group(e.g. alkyl) is indicated by the prefix “C_(x)-C_(y)” or “C_(x-y)”,wherein x is the minimum and y is the maximum number of carbon atoms inthe group. Thus, for example, “(C₁-C₆)alkyl” or “C₁₋₆ alkyl” refers toan alkyl substituent containing from 1 to 6 carbon atoms.

The term “halogen”, as used herein, refers to fluoride, chloride,bromide, or iodide.

The term “haloalkyl”, as used herein, refers to an alkyl group that issubstituted with at least one halogen substituent. Where more than onehydrogen atom is replaced with a halogen atom, the halogens may beidentical or different. Non-limiting examples include fluoromethyl,difluoromethyl, trifluoromethyl and 2,2,2-trifluoroethyl.

As used herein, a “biotherapeutic agent” means a biological molecule,such as an antibody or fusion protein, that blocks ligand/receptorsignaling in any biological pathway that supports tumor maintenanceand/or growth or suppresses the anti-tumor immune response.

As used herein, a “chemotherapeutic agent” is a chemical compound usefulin the treatment of cancer. Chemotherapeutic agents useful in thetreatment methods of the present invention include cytostatic and/orcytotoxic agents. Chemotherapeutic agents include the agent itself orany pharmaceutically acceptable salt, co-crystal, or solvate thereof.Chemotherapeutic agents are further described herein.

As used herein, the following terms are used interchangeably and meanany one or more therapeutic agent, other than a compound of theinvention, that is or can be used in the treatment of cancer:“additional anti-cancer therapeutic agent” or “additionalchemotherapeutic agent” or “additional therapeutic agent.”

A biotherapeutic agent and a chemotherapeutic agent are both examples ofan additional anti-cancer therapeutic agent.

As used herein, a “cytotoxic agent” refers to an agent that has acytotoxic and/or cytostatic effect on a cell and a “cytostatic effect”refers to the inhibition of cell proliferation.

As used herein, a “cytostatic agent” refers to an agent that has acytostatic effect on a cell, thereby inhibiting the growth and/orexpansion of a specific subset of cells (i.e., tumor cells).

As used herein, an “immunomodulating agent” refers to an agent thatstimulates the immune response though the production of cytokines and/orantibodies and/or modulating T cell function thereby inhibiting orreducing the growth of a subset of cells (i.e., tumor cells) eitherdirectly or indirectly by allowing another agent to be more efficacious.

“Consists essentially of,” and variations such as “consist essentiallyof” or “consisting essentially of,” as used throughout the specificationand claims, indicate the inclusion of any recited elements or group ofelements, and the optional inclusion of other elements, of similar ordifferent nature than the recited elements, that do not materiallychange the basic or novel properties of the specified dosage regimen,method, or composition. As a non-limiting example, an OX40 agonist thatconsists essentially of a recited amino acid sequence may also includeone or more amino acids, including substitutions of one or more aminoacid residues, which do not materially affect the properties of thebinding compound.

As used herein, an “effective dosage” or “effective amount” of drug,compound or pharmaceutical composition is an amount sufficient to affectany one or more beneficial or desired, including biochemical,histological and/or behavioral symptoms, of the disease, itscomplications and intermediate pathological phenotypes presenting duringdevelopment of the disease. For therapeutic use, a “therapeuticallyeffective amount” refers to that amount of a compound being administeredwhich will relieve to some extent one or more of the symptoms of thedisorder being treated. In reference to the treatment of cancer, atherapeutically effective amount refers to that amount which has theeffect of (1) reducing the size of the tumor, (2) inhibiting (that is,slowing to some extent, preferably stopping) tumor metastasis, (3)inhibiting to some extent (that is, slowing to some extent, preferablystopping) tumor growth or tumor invasiveness, (4) relieving to someextent (or, preferably, eliminating) one or more signs or symptomsassociated with the cancer, (5) decreasing the dose of other medicationsrequired to treat the disease, and/or (6) enhancing the effect ofanother medication, and/or (7) delaying the progression of the diseasein a patient.

An effective dosage can be administered in one or more administrations.For the purposes of this invention, an effective dosage of drug,compound, or pharmaceutical composition is an amount sufficient toaccomplish prophylactic or therapeutic treatment either directly orindirectly. As is understood in the clinical context, an effectivedosage of drug, compound or pharmaceutical composition may or may not beachieved in conjunction with another drug, compound or pharmaceuticalcomposition.

A “pharmaceutical composition” refers to a mixture of one or more of thecompounds of the invention, or a pharmaceutically acceptable salt,solvate, hydrate or prodrug thereof as an active ingredient, and atleast one pharmaceutically acceptable carrier or excipient. In someembodiments, the pharmaceutical composition comprises two or morepharmaceutically acceptable carriers and/or excipients. In otherembodiments, the pharmaceutical composition further comprises at leastone additional anticancer therapeutic agent.

In one aspect, the invention provides a pharmaceutical compositioncomprising a compound of the invention, or a pharmaceutically acceptablesalt thereof, and a pharmaceutically acceptable carrier or excipient. Insome embodiments, the pharmaceutical composition comprises two or morepharmaceutically acceptable carriers and/or excipients.

In some embodiments, the pharmaceutical composition further comprises atleast one additional anti-cancer therapeutic agent. In some suchembodiments, the combination provides an additive, greater thanadditive, or synergistic anti-cancer effect.

“Tumor” as it applies to a subject diagnosed with, or suspected ofhaving, a cancer refers to a malignant or potentially malignant neoplasmor tissue mass of any size and includes primary tumors and secondaryneoplasms. A solid tumor is an abnormal growth or mass of tissue thatusually does not contain cysts or liquid areas. Examples of solid tumorsare sarcomas, carcinomas, and lymphomas. Leukemias (cancers of theblood) generally do not form solid tumors (National Cancer Institute,Dictionary of Cancer Terms).

“Tumor burden” or “tumor load’, refers to the total amount of tumorousmaterial distributed throughout the body. Tumor burden refers to thetotal number of cancer cells or the total size of tumor(s), throughoutthe body, including lymph nodes and bone marrow. Tumor burden can bedetermined by a variety of methods known in the art, such as, e.g.,using calipers, or while in the body using imaging techniques, e.g.,ultrasound, bone scan, computed tomography (CT), or magnetic resonanceimaging (MRI) scans.

The term “tumor size” refers to the total size of the tumor which can bemeasured as the length and width of a tumor. Tumor size may bedetermined by a variety of methods known in the art, such as, e.g., bymeasuring the dimensions of tumor(s) upon removal from the subject,e.g., using calipers, or while in the body using imaging techniques,e.g., bone scan, ultrasound, CR or MRI scans.

As used herein, “subject” refers to a human or animal subject. When thesubject is a human, the subject may also be referred to as a “patient”.

The term “treat” or “treating” a cancer as used herein means toadminister a compound of the present invention to a subject havingcancer, or diagnosed with cancer, to achieve at least one positivetherapeutic effect, such as, for example, reduced number of cancercells, reduced tumor size, reduced rate of cancer cell infiltration intoperipheral organs, or reduced rate of tumor metastases or tumor growth,reversing, alleviating, inhibiting the progress of, or preventing thedisorder or condition to which such term applies, or one or moresymptoms of such disorder or condition. The term “treatment”, as usedherein, unless otherwise indicated, refers to the act of treating as“treating” is defined immediately above. The term “treating” alsoincludes adjuvant and neo-adjuvant treatment of a subject.

For the purposes of this invention, beneficial or desired clinicalresults include, but are not limited to, one or more of the following:reducing the proliferation of (or destroying) neoplastic or cancerouscell; inhibiting metastasis or neoplastic cells; shrinking or decreasingthe size of a tumor; remission of the cancer; decreasing symptomsresulting from the cancer; increasing the quality of life of thosesuffering from the cancer; decreasing the dose of other medicationsrequired to treat the cancer; delaying the progression of the cancer;curing the cancer; overcoming one or more resistance mechanisms of thecancer; and/or prolonging survival of patients the cancer. Positivetherapeutic effects in cancer can be measured in several ways (see, forexample, W. A. Weber, Assessing tumor response to therapy, J. Nucl. Med.50 Suppl. 1:1S-10S (2009). For example, with respect to tumor growthinhibition (T/C), according to the National Cancer Institute (NCI)standards, a T/C less than or equal to 42% is the minimum level ofanti-tumor activity. A T/C<10% is considered a high anti-tumor activitylevel, with T/C (%)=median tumor volume of the treated/median tumorvolume of the control×100.

In some embodiments, the treatment achieved by a compound of theinvention is defined by reference to any of the following: partialresponse (PR), complete response (CR), overall response (OR),progression free survival (PFS), disease free survival (DFS) and overallsurvival (OS). PFS, also referred to as “Time to Tumor Progression”indicates the length of time during and after treatment that the cancerdoes not grow and includes the amount of time patients have experienceda CR or PR, as well as the amount of time patients have experiencedstable disease (SD). DFS refers to the length of time during and aftertreatment that the patient remains free of disease. OS refers to aprolongation in life expectancy as compared to naïve or untreatedsubjects or patients. In some embodiments, response to a combination ofthe invention is any of PR, CR, PFS, DFS, OR or OS that is assessedusing Response Evaluation Criteria in Solid Tumors (RECIST) 1.1 responsecriteria.

The term “additive” is used to mean that the result of the combinationof two compounds, components or targeted agents is no greater than thesum of each compound, component or targeted agent individually.

The term “synergy” or “synergistic” are used to mean that the result ofthe combination of two compounds, components or targeted agents isgreater than the sum of each compound, component or targeted agentindividually. This improvement in the disease, condition or disorderbeing treated is a “synergistic” effect. A “synergistic amount” is anamount of the combination of the two compounds, components or targetedagents that results in a synergistic effect, as “synergistic” is definedherein.

Determining a synergistic interaction between one or two components, theoptimum range for the effect and absolute dose ranges of each componentfor the effect may be definitively measured by administration of thecomponents over different dose ranges, and/or dose ratios to patients inneed of treatment. However, the observation of synergy in in vitromodels or in vivo models can be predictive of the effect in humans andother species and in vitro models or in vivo models exist, as describedherein, to measure a synergistic effect. The results of such studies canalso be used to predict effective dose and plasma concentration ratioranges and the absolute doses and plasma concentrations required inhumans and other species such as by the application of pharmacokineticand/or pharmacodynamics methods.

The treatment regimen for a compound of the invention that is effectiveto treat a cancer patient may vary according to factors such as thedisease state, age, and weight of the patient, and the ability of thetherapy to elicit an anti-cancer response in the subject. While anembodiment of any of the aspects of the invention may not be effectivein achieving a positive therapeutic effect in every subject, it shoulddo so in a statistically significant number of subjects as determined byany statistical test known in the art such as the Student's t-test, thechi2-test the U-test according to Mann and Whitney, the Kruskal-Wallistest (H-test), Jonckheere-Terpstrat-testy and the Wilcon on-test.

The terms “treatment regimen”, “dosing protocol” and “dosing regimen”are used interchangeably to refer to the dose and timing ofadministration of each compound of the invention, alone or incombination with another therapeutic agent.

“Ameliorating” means a lessening or improvement of one or more symptomsupon treatment with a combination described herein, as compared to notadministering the combination. “Ameliorating” also includes shorteningor reduction in duration of a symptom.

“Abnormal cell growth”, as used herein, unless otherwise indicated,refers to cell growth that is independent of normal regulatorymechanisms (e.g., loss of contact inhibition). Abnormal cell growth maybe benign (not cancerous), or malignant (cancerous).

The terms “cancer” or “cancerous” refers to any malignant and/orinvasive growth or tumor caused by abnormal cell growth. Cancer includesprimary cancer that originates at a specific site in the body, ametastatic cancer that has spread from the place in which it started toother parts of the body, a recurrence from the original primary cancerafter remission, and a second primary cancer that is a new primarycancer in a patient with a history of previous cancer of a differenttype from the second primary cancer. Cancer includes solid tumors namedfor the type of cells that form them, cancer of blood, bone marrow, orthe lymphatic system. Examples of solid tumors include sarcomas andcarcinomas. Cancers of the blood include leukemia, lymphoma and myeloma.Additional examples of cancer include blastomas and an actinickeratosis. Cancer also includes primary cancer or metastases of a siteselected from the group consisting of oral cavity, digestive system,respiratory system, skin, breast, genital system, urinary system, ocularsystem, nervous system, endocrine system, and lymphoma.

Unless indicated otherwise, all references herein to the inventivecompounds include references to salts, solvates, hydrates and complexesthereof, and to solvates, hydrates and complexes of salts thereof,including polymorphs, stereoisomers, and isotopically labelled versionsthereof.

Compounds of the invention may exist in the form of pharmaceuticallyacceptable salts such as, e.g., acid addition salts and base additionsalts of the compounds of one of the formulae provided herein. As usedherein, the term “pharmaceutically acceptable salt” refers to thosesalts which retain the biological effectiveness and properties of theparent compound. The phrase “pharmaceutically acceptable salt(s)”, asused herein, unless otherwise indicated, includes salts of acidic orbasic groups which may be present in the compounds of the formulaedisclosed herein.

For example, the compounds of the invention that are basic in nature arecapable of forming a wide variety of salts with various inorganic andorganic acids. Although such salts must be pharmaceutically acceptablefor administration to animals, it is often desirable in practice toinitially isolate the compound of the present invention from thereaction mixture as a pharmaceutically unacceptable salt and then simplyconvert the latter back to the free base compound by treatment with analkaline reagent and subsequently convert the latter free base to apharmaceutically acceptable acid addition salt. The acid addition saltsof the base compounds of this invention can be prepared by treating thebase compound with a substantially equivalent amount of the selectedmineral or organic acid in an aqueous solvent medium or in a suitableorganic solvent, such as methanol or ethanol. Upon evaporation of thesolvent, the desired solid salt is obtained. The desired acid salt canalso be precipitated from a solution of the free base in an organicsolvent by adding an appropriate mineral or organic acid to thesolution.

The acids that may be used to prepare pharmaceutically acceptable acidaddition salts of such basic compounds of those that form non toxic acidaddition salts, i.e., salts containing pharmacologically acceptableanions, such as the hydrochloride, hydrobromide, hydroiodide, nitrate,sulfate, bisulfate, phosphate, acid phosphate, isonicotinate, acetate,lactate, salicylate, citrate, acid citrate, tartrate, pantothenate,bitartrate, ascorbate, succinate, maleate, gentisinate, fumarate,gluconate, glucuronate, saccharate, formate, benzoate, glutamate,methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonateand pamoate salts.

Examples of salts include, but are not limited to, acetate, acrylate,benzenesulfonate, benzoate (such as chlorobenzoate, methylbenzoate,dinitrobenzoate, hydroxybenzoate, and methoxybenzoate), bicarbonate,bisulfate, bisulfite, bitartrate, borate, bromide, butyne 1,4 dioate,calcium edetate, camsylate, carbonate, chloride, caproate, caprylate,clavulanate, citrate, decanoate, dihydrochloride, dihydrogenphosphate,edetate, edislyate, estolate, esylate, ethylsuccinate, formate,fumarate, gluceptate, gluconate, glutamate, glycollate,glycollylarsanilate, heptanoate, hexyne 1,6 dioate, hexylresorcinate,hydrabamine, hydrobromide, hydrochloride, y hydroxybutyrate, iodide,isobutyrate, isothionate, lactate, lactobionate, laurate, malate,maleate, malonate, mandelate, mesylate, metaphosphate, methanesulfonate, methylsulfate, monohydrogenphosphate, mucate, napsylate,naphthalene 1 sulfonate, naphthalene 2 sulfonate, nitrate, oleate,oxalate, pamoate (embonate), palmitate, pantothenate, phenylacetates,phenylbutyrate, phenylpropionate, phthalate, phospate/diphosphate,polygalacturonate, propanesulfonate, propionate, propiolate,pyrophosphate, pyrosulfate, salicylate, stearate, subacetate, suberate,succinate, sulfate, sulfonate, sulfite, tannate, tartrate, teoclate,tosylate and valerate salts.

Illustrative examples of suitable salts include organic salts derivedfrom amino acids, such as glycine and arginine, ammonia, primary,secondary, and tertiary amines and cyclic amines, such as piperidine,morpholine and piperazine, and inorganic salts derived from sodium,calcium, potassium, magnesium, manganese, iron, copper, zinc, aluminumand lithium.

The compounds of the invention that include a basic moiety, such as anamino group, may form pharmaceutically acceptable salts with variousamino acids, in addition to the acids mentioned above.

Alternatively, the compounds useful that are acidic in nature may becapable of forming base salts with various pharmacologically acceptablecations. Examples of such salts include the alkali metal or alkalineearth metal salts and particularly, the sodium and potassium salts.These salts are all prepared by conventional techniques. The chemicalbases which are used as reagents to prepare the pharmaceuticallyacceptable base salts of this invention are those which form non toxicbase salts with the acidic compounds herein. These salts may be preparedby any suitable method, for example, treatment of the free acid with aninorganic or organic base, such as an amine (primary, secondary ortertiary), an alkali metal hydroxide or alkaline earth metal hydroxide,or the like. These salts can also be prepared by treating thecorresponding acidic compounds with an aqueous solution containing thedesired pharmacologically acceptable cations, and then evaporating theresulting solution to dryness, preferably under reduced pressure.Alternatively, they may also be prepared by mixing lower alkanolicsolutions of the acidic compounds and the desired alkali metal alkoxidetogether, and then evaporating the resulting solution to dryness in thesame manner as before. In either case, stoichiometric quantities ofreagents are preferably employed in order to ensure completeness ofreaction and maximum yields of the desired final product.

The chemical bases that may be used as reagents to preparepharmaceutically acceptable base salts of the compounds of the inventionthat are acidic in nature are those that form non toxic base salts withsuch compounds. Such non toxic base salts include, but are not limitedto, those derived from such pharmacologically acceptable cations such asalkali metal cations (e.g., potassium and sodium) and alkaline earthmetal cations (e.g., calcium and magnesium), ammonium or water solubleamine addition salts such as N methylglucamine (meglumine), and thelower alkanolammonium and other base salts of pharmaceuticallyacceptable organic amines.

Hemisalts of acids and bases may also be formed, for example,hemisulphate and hemicalcium salts.

For a review on suitable salts, see Handbook of Pharmaceutical Salts:Properties, Selection, and Use by Stahl and Wermuth (Wiley VCH, 2002).Methods for making pharmaceutically acceptable salts of compounds of theinvention, and of interconverting salt and free base forms, are known toone of skill in the art.

Salts of the present invention can be prepared according to methodsknown to those of skill in the art. A pharmaceutically acceptable saltof the inventive compounds can be readily prepared by mixing togethersolutions of the compound and the desired acid or base, as appropriate.The salt may precipitate from solution and be collected by filtration ormay be recovered by evaporation of the solvent. The degree of ionizationin the salt may vary from completely ionized to almost non ionized.

It will be understood by those of skill in the art that the compounds ofthe invention in free base form having a basic functionality may beconverted to the acid addition salts by treating with a stoichiometricexcess of the appropriate acid. The acid addition salts of the compoundsof the invention may be reconverted to the corresponding free base bytreating with a stoichiometric excess of a suitable base, such aspotassium carbonate or sodium hydroxide, typically in the presence ofaqueous solvent, and at a temperature of between about 0° C. and 100° C.The free base form may be isolated by conventional means, such asextraction with an organic solvent. In addition, acid addition salts ofthe compounds of the invention may be interchanged by taking advantageof differential solubilities of the salts, volatilities or acidities ofthe acids, or by treating with the appropriately loaded ion exchangeresin. For example, the interchange may be affected by the reaction of asalt of the compounds of the invention with a slight stoichiometricexcess of an acid of a lower pK than the acid component of the startingsalt. This conversion is typically carried out at a temperature betweenabout 0° C. and the boiling point of the solvent being used as themedium for the procedure. Similar exchanges are possible with baseaddition salts, typically via the intermediacy of the free base form.

The compounds of the invention may exist in both unsolvated and solvatedforms. When the solvent or water is tightly bound, the complex will havea well defined stoichiometry independent of humidity. When, however, thesolvent or water is weakly bound, as in channel solvates and hygroscopiccompounds, the water/solvent content will be dependent on humidity anddrying conditions. In such cases, non stoichiometry will be the norm.The term ‘solvate’ is used herein to describe a molecular complexcomprising the compound of the invention and one or morepharmaceutically acceptable solvent molecules, for example, ethanol. Theterm ‘hydrate’ is employed when the solvent is water. Pharmaceuticallyacceptable solvates in accordance with the invention include hydratesand solvates wherein the solvent of crystallization may be isotopicallysubstituted, e.g. D₂O, d₆-acetone, d₆-DMSO.

Also included within the scope of the invention are complexes such asclathrates, drug host inclusion complexes wherein, in contrast to theaforementioned solvates, the drug and host are present in stoichiometricor non stoichiometric amounts. Also included are complexes of the drugcontaining two or more organic and/or inorganic components which may bein stoichiometric or non stoichiometric amounts. The resulting complexesmay be ionized, partially ionized, or non ionized. For a review of suchcomplexes, see J Pharm Sci, 64 (8), 1269 1288 by Haleblian (August1975), the disclosure of which is incorporated herein by reference inits entirety.

The invention also relates to prodrugs of the compounds of the formulaeprovided herein. Thus, certain derivatives of compounds of the inventionwhich may have little or no pharmacological activity themselves can,when administered to a patient, be converted into the inventivecompounds, for example, by hydrolytic cleavage. Such derivatives arereferred to as ‘prodrugs’. Further information on the use of prodrugsmay be found in ‘Pro drugs as Novel Delivery Systems, Vol. 14, ACSSymposium Series (T Higuchi and W Stella); ‘Bioreversible Carriers inDrug Design’, Pergamon Press, 1987 (ed. E B Roche, AmericanPharmaceutical Association), and Guarino, V. R; Stella, V. J.: BiotechPharm. Aspects 2007 5 (Pt2) 133-187, the disclosures of which areincorporated herein by reference in their entireties.

Prodrugs in accordance with the invention can, for example, be producedby replacing appropriate functionalities present in the inventivecompounds with certain moieties known to those skilled in the art as‘pro moieties’ as described, for example, in “Design of Prodrugs” by HBundgaard (Elsevier, 1985), the disclosure of which is incorporatedherein by reference in its entirety.

Some non limiting examples of prodrugs in accordance with the inventioninclude:

(i) where the compound contains a carboxylic acid functionality (COOH),an ester thereof, for example, replacement of the hydrogen with(C₁-C₆)alkyl;

(ii) where the compound contains an alcohol functionality (OH), an etherthereof, for example, replacement of the hydrogen with(C₁-C₆)alkanoyloxymethyl, or with a phosphate ether group; and

(iii) where the compound contains a primary or secondary aminofunctionality (NH2 or NHR where R≠H), an amide thereof, for example,replacement of one or both hydrogens with a suitably metabolicallylabile group, such as an amide, carbamate, urea, phosphonate, sulfonate,etc.

Further examples of replacement groups in accordance with the foregoingexamples and examples of other prodrug types may be found in theaforementioned references.

Finally, certain inventive compounds may themselves act as prodrugs ofother of the inventive compounds.

Also included within the scope of the invention are metabolites ofcompounds of the formulae described herein, i.e., compounds formed invivo upon administration of the drug.

The compounds of the formulae provided herein may have asymmetric carbonatoms. The carbon carbon bonds of the compounds of the invention may bedepicted herein using a solid line (

), a solid wedge (

), or a dotted wedge (

). The use of a solid line to depict bonds to asymmetric carbon atoms ismeant to indicate that all possible stereoisomers (e.g. specificenantiomers, racemic mixtures, etc.) at that carbon atom are included.The use of either a solid or dotted wedge to depict bonds to asymmetriccarbon atoms is meant to indicate that only the stereoisomer shown ismeant to be included. It is possible that compounds of the invention maycontain more than one asymmetric carbon atom. In those compounds, theuse of a solid line to depict bonds to asymmetric carbon atoms is meantto indicate that all possible stereoisomers are meant to be included andthe attached stereocenter. For example, unless stated otherwise, it isintended that the compounds of the invention can exist as enantiomersand diastereomers or as racemates and mixtures thereof. The use of asolid line to depict bonds to one or more asymmetric carbon atoms in acompound of the invention and the use of a solid or dotted wedge todepict bonds to other asymmetric carbon atoms in the same compound ismeant to indicate that a mixture of diastereomers is present.

Compounds of the invention that have chiral centers may exist asstereoisomers, such as racemates, enantiomers, or diastereomers.

Stereoisomers of the compounds of the formulae herein can include cisand trans isomers, optical isomers such as (R) and (S) enantiomers,diastereomers, geometric isomers, rotational isomers, atropisomers,conformational isomers, and tautomers of the compounds of the invention,including compounds exhibiting more than one type of isomerism; andmixtures thereof (such as racemates and diastereomeric pairs).

Also included are acid addition salts or base addition salts, whereinthe counterion is optically active, for example, d lactate or I lysine,or racemic, for example, dl tartrate or dl arginine.

When any racemate crystallizes, crystals of two different types arepossible. The first type is the racemic compound (true racemate)referred to above wherein one homogeneous form of crystal is producedcontaining both enantiomers in equimolar amounts. The second type is theracemic mixture or conglomerate wherein two forms of crystal areproduced in equimolar amounts each comprising a single enantiomer.

The compounds of the invention may exhibit the phenomena of tautomerismand structural isomerism. For example, the compounds may exist inseveral tautomeric forms, including the enol and imine form, and theketo and enamine form and geometric isomers and mixtures thereof. Allsuch tautomeric forms are included within the scope of compounds of theinvention. Tautomers exist as mixtures of a tautomeric set in solution.In solid form, usually one tautomer predominates. Even though onetautomer may be described, the present invention includes all tautomersof the compounds of the formulae provided.

In addition, some of the compounds of the invention may formatropisomers (e.g., substituted biaryls). Atropisomers areconformational stereoisomers which occur when rotation about a singlebond in the molecule is prevented, or greatly slowed, as a result ofsteric interactions with other parts of the molecule and thesubstituents at both ends of the single bond are unsymmetrical. Theinterconversion of atropisomers is slow enough to allow separation andisolation under predetermined conditions. The energy barrier to thermalracemization may be determined by the steric hindrance to free rotationof one or more bonds forming a chiral axis.

Where a compound of the invention contains an alkenyl or alkenylenegroup, geometric cis/trans (or Z/E) isomers are possible. Cis/transisomers may be separated by conventional techniques well known to thoseskilled in the art, for example, chromatography and fractionalcrystallization.

Conventional techniques for the preparation/isolation of individualenantiomers include chiral synthesis from a suitable optically pureprecursor or resolution of the racemate (or the racemate of a salt orderivative) using, for example, chiral high-pressure liquidchromatography (HPLC) or superfluid critical chromatography (SFC).

Alternatively, the racemate (or a racemic precursor) may be reacted witha suitable optically active compound, for example, an alcohol, or, inthe case where the compound contains an acidic or basic moiety, an acidor base such as tartaric acid or 1 phenylethylamine. The resultingdiastereomeric mixture may be separated by chromatography and/orfractional crystallization and one or both of the diastereoisomersconverted to the corresponding pure enantiomer(s) by means well known toone skilled in the art.

Chiral compounds of the invention (and chiral precursors thereof) may beobtained in enantiomerically enriched form using chromatography,typically HPLC, on an asymmetric resin with a mobile phase consisting ofa hydrocarbon, typically heptane or hexane, containing from 0 to 50%isopropanol, typically from 2 to 20%, and from 0 to 5% of an alkylamine,typically 0.1% diethylamine. Concentration of the eluate affords theenriched mixture.

Stereoisomeric conglomerates may be separated by conventional techniquesknown to those skilled in the art; see, for example, “Stereochemistry ofOrganic Compounds” by E L Eliel (Wiley, New York, 1994), the disclosureof which is incorporated herein by reference in its entirety.

The enantiomeric purity of compounds described herein may be describedin terms of enantiomeric excess (ee), which indicates the degree towhich a sample contains one enantiomer in greater amounts than theother. A racemic mixture has an ee of 0%, while a single completely pureenantiomer has an ee of 100%. Similarly, diastereomeric purity may bedescribed in terms of diasteriomeric excess (de).

The present invention also includes isotopically labeled compounds,which are identical to those recited in one of the formulae provided,but for the fact that one or more atoms are replaced by an atom havingan atomic mass or mass number different from the atomic mass or massnumber usually found in nature.

Isotopically labeled compounds of the invention can generally beprepared by conventional techniques known to those skilled in the art orby processes analogous to those described herein, using an appropriateisotopically labeled reagent in place of the non labeled reagentotherwise employed.

Examples of isotopes that may be incorporated into compounds of theinvention include isotopes of hydrogen, carbon, nitrogen, oxygen,phosphorus, sulfur, fluorine and chlorine, such as, but not limited to,²H, ³H, ¹³C, ¹⁴C, ¹⁵N, ¹⁸O, ¹⁷O, ³²P, ³⁵S, ¹⁸F and ³⁶Cl. Certainisotopically labeled compounds of the invention, for example those intowhich radioactive isotopes such as ³H and ¹⁴C are incorporated, areuseful in drug and/or substrate tissue distribution assays. Tritiated,i.e., ³H, and carbon 14, i.e., ¹⁴C, isotopes are particularly preferredfor their ease of preparation and detectability. Further, substitutionwith heavier isotopes such as deuterium, i.e., ²H, can afford certaintherapeutic advantages resulting from greater metabolic stability, forexample increased in vivo half life or reduced dosage requirements and,hence, may be preferred in some circumstances. Isotopically labeledcompounds of the invention may generally be prepared by carrying out theprocedures disclosed in the Schemes and/or in the Examples andPreparations below, by substituting an isotopically labeled reagent fora non isotopically labeled reagent.

Compounds of the invention intended for pharmaceutical use may beadministered as crystalline or amorphous products, or mixtures thereof.They may be obtained, for example, as solid plugs, powders, or films bymethods such as precipitation, crystallization, freeze drying, spraydrying, or evaporative drying. Microwave or radio frequency drying maybe used.

In another embodiment, the invention provides a compound of Formula (Ia)

or a pharmaceutically acceptable salt thereof, wherein

R¹ and R² are independently C₁₋₂ alkyl; or

R¹ and R² are joined to form a 5- to 7-membered carbocyclic ring,wherein said carbocyclic ring may be saturated or unsaturated;

R³ is

R⁴ is C₃₋₅ alkyl, or (CH₂)_(n)O(CH₂)_(m)CH₃;

R⁵ is C₁₋₂ alkyl;

m is 1; and

n is 1.

In another embodiment, the invention provides a compound of Formula(Ia), or a pharmaceutically acceptable salt thereof, wherein R¹ and R²are independently C₁₋₂ alkyl.

In another embodiment, the invention provides a compound of Formula(Ia), or a pharmaceutically acceptable salt thereof, wherein R¹ and R²are joined to form a 5- to 7-membered carbocyclic ring, wherein saidcarbocyclic ring may be saturated or unsaturated.

In another embodiment, the invention provides a compound of Formula(Ia), or a pharmaceutically acceptable salt thereof, wherein

R¹ and R² are joined to form a 5- to 7-membered carbocyclic ring,wherein said carbocyclic ring may be saturated; and

R⁴ is (CH₂)_(n)O(CH₂)_(m)CH₃.

In another embodiment, the invention provides a compound of Formula(Ia), or a pharmaceutically acceptable salt thereof, wherein thecarbocyclic ring is cyclopentyl.

In another embodiment, the invention provides a compound of Formula(Ia), or a pharmaceutically acceptable salt thereof, wherein thecarbocyclic ring is cyclohexyl.

In another embodiment, the invention provides a compound of Formula(Ia), or a pharmaceutically acceptable salt thereof, wherein

R¹ and R² are joined to form a 5- to 7-membered carbocyclic ring,wherein said carbocyclic ring may be unsaturated; and

R⁴ is C₃₋₅ alkyl.

In another embodiment, the invention provides a compound of Formula(Ia), or a pharmaceutically acceptable salt thereof, wherein thecarbocyclic ring is phenyl.

In another embodiment, the invention provides a compound of Formula (Ib)

or a pharmaceutically acceptable salt thereof, wherein

R¹ and R² are independently C₁₋₃ alkyl; or

R¹ and R² are joined to form a 5- to 7-membered carbocyclic ring,wherein said carbocyclic ring may be saturated or unsaturated;

R³ is

R⁴ is C₁₋₆ alkyl, or (CH₂)_(n)O(CH₂)_(m)CH₃, wherein said C₁₋₆ alkyl orany carbon of the (CH₂)_(n)O(CH₂)_(m)CH₃ group is substituted with 0 to3 halogen as valency allows, wherein halogen is F;

R⁵ is C₁₋₃ alkyl, or OC₁₋₃ alkyl, wherein the C₁₋₃ alkyl is substitutedby 0 to 3 F;

R⁶ is H, or C₁₋₃ alkyl, wherein the C₁₋₃ alkyl is substituted with 0 to3 F;

m is 0 to 2; and

n is 1 to 3.

In another embodiment, the invention provides a compound of Formula(Ib), or a pharmaceutically acceptable salt thereof, wherein

R¹ and R² are independently C₁₋₂ alkyl; or

R¹ and R² are joined to form a 5- to 7-membered carbocyclic ring,wherein said carbocyclic ring may be saturated or unsaturated;

R³ is

R⁵ is C₁₋₃ alkyl, or OC₁₋₃ alkyl, wherein the C₁₋₃ alkyl is substitutedby 0 to 2 F; and

R⁶ is H.

In another embodiment, the invention provides a compound of Formula(Ib), or a pharmaceutically acceptable salt thereof, wherein R⁵ is C₁₋₂alkyl.

In another embodiment, the invention provides a compound of Formula (I),(Ia), (Ib), or a pharmaceutically acceptable salt thereof, wherein R⁴ isn-propyl, n-butyl, or n-pentyl.

In another embodiment, the invention provides a compound of Formula (I),(Ia), (Ib), or a pharmaceutically acceptable salt thereof, wherein R⁴ is—CH₂—O—CH₂CH₃.

In another embodiment, the invention provides a compound of Formula (I),(Ia), (Ib), or a pharmaceutically acceptable salt thereof, wherein R⁵ ismethyl or ethyl.

In another embodiment, the invention provides a compound of Formula (I),(Ia), (Ib), or a pharmaceutically acceptable salt thereof, wherein R⁶ isH.

Another embodiment of the invention is one or more of each Exampledescribed herein, and includes, but is not limited to, the compoundsselected from:

-   2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol;-   2-((4-amino-2-(ethoxymethyl)-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;-   2-((4-amino-2-(ethoxymethyl)-7,8-dihydrocyclopenta[b]imidazo[4,5-d]pyridin-1(6H)-yl)methyl)-2-methylpropane-1,3-diol;-   2-((4-amino-2-(ethoxymethyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;-   3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-ol;-   (R)-3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-ol;-   (S)-3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-ol;-   2-((4-amino-6,7-dimethyl-2-(2,2,2-trifluoroethyl)-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol;-   2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-ethylpropane-1,3-diol;-   2-((4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;-   2-((4-amino-2-butyl-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol;    and-   2-((4-amino-2-pentyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;    or a pharmaceutically acceptable salt thereof.

Another embodiment of the invention is one or more of each Exampledescribed herein, and includes, but is not limited to, the compoundsselected from:

-   2-((4-amino-2-butyl-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;-   2-((4-amino-2-butyl-7,8-dihydrocyclopenta[b]imidazo[4,5-d]pyridin-1(6H)-yl)methyl)-2-methylpropane-1,3-diol;-   2-((4-amino-2-ethyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;    and-   2-((4-amino-2-propyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;    or a pharmaceutically acceptable salt thereof.

Another embodiment of the invention provides a pharmaceuticalcomposition comprising the compounds of Formula (I) and any embodimentsthereof, or a pharmaceutically acceptable salt thereof, and apharmaceutically acceptable carrier or excipient.

Another embodiment of the invention provides a method for the treatmentof cancer in a subject in need thereof, comprising administering to thesubject a therapeutically effective amount of the compounds of Formula(I) and any embodiments thereof, or a pharmaceutically acceptable saltthereof.

Cancers to be treated include squamous cell carcinoma, basal cellcarcinomas, myeloma, small-cell lung cancer, non-small cell lung cancer,glioma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, acute myeloidleukemia (AML), multiple myeloma, gastrointestinal (tract) cancer, renalcancer, ovarian cancer, liver cancer, lymphoblastic leukemia,lymphocytic leukemia, colorectal cancer, endometrial cancer, kidneycancer, prostate cancer, thyroid cancer, melanoma, chondrosarcoma,neuroblastoma, pancreatic cancer, glioblastoma multiforme, cervicalcancer, brain cancer, stomach cancer, uterine cancer, bladder cancer,including non-muscular invasive bladder cancer, hepatoma, breast cancer,and head and neck cancer.

More particular examples of cancers to be treated include basal cellcarcinomas, small-cell lung cancer, non-small cell lung cancer,non-Hodgkin's lymphoma, ovarian cancer, colorectal cancer, kidneycancer, prostate cancer, thyroid cancer, melanoma, pancreatic cancer,bladder cancer (non-muscular invasive bladder cancer), hepatoma, breastcancer, and head and neck cancer.

Another embodiment of the invention concerns treatment of cancersselected from basal cell carcinomas, ovarian cancer, melanoma,non-muscular invasive bladder cancer, breast cancer, and head and neckcancer.

Another embodiment of the invention concerns treatment melanoma,gastrointestinal (tract) cancer, breast cancer, ovarian cancer, and headand neck cancer.

Another embodiment of the invention concerns treatment cancers of thegastrointestinal tract. Such gastrointestinal cancers include cancer ofthe mouth, esophagus, stomach, biliary system, pancreas, smallintestine, large intestine, rectum, and anus.

Another embodiment of the invention concerns treatment of non-muscularinvasive bladder cancer.

Another embodiment of the invention concerns the compounds of Formula(I) and any embodiments thereof, or a pharmaceutically acceptable saltthereof, for use in the treatment of cancer in a subject in needthereof.

Another embodiment of the invention concerns the compounds of Formula(I) and any embodiments thereof, or a pharmaceutically acceptable saltthereof, for use in the treatment of cancer, wherein said treatmentcomprises the administration of an additional therapeutic agent.

In another aspect, the invention provides a method of inhibiting cancercell proliferation in a subject, comprising administering to the subjecta compound of the invention, or a pharmaceutically acceptable saltthereof, in an amount effective to inhibit cell proliferation.

In another aspect, the invention provides a method of inhibiting cancercell invasiveness in a subject, comprising administering to the subjecta compound of the invention, or a pharmaceutically acceptable saltthereof, in an amount effective to inhibit cell invasiveness.

In another aspect, the invention provides a method of inducing apoptosisin cancer cells in a subject, comprising administering to the subject acompound of the invention, or a pharmaceutically acceptable saltthereof, in an amount effective to induce apoptosis.

In another aspect, the invention provides a method of inhibiting cancercell metastasis in a subject, comprising administering to the subject acompound of the invention, or a pharmaceutically acceptable saltthereof, in an amount effective to inhibit cell metastasis.

In another aspect, the invention provides a method of inhibitingangiogenesis in a subject, comprising administering to the subject acompound of the invention, or a pharmaceutically acceptable saltthereof, in an amount effective to inhibit angiogenesis.

In another aspect, the invention provides a method for the treatment ofcancer in a subject in need thereof, comprising administering to thesubject a therapeutically effective amount of the compound of theinvention, or a pharmaceutically acceptable salt thereof. Said methodalso includes administering the compound of the invention with at leastone additional therapeutic agent.

The invention also provides methods of preventing an infectious diseasein a subject in need thereof, comprising administration of apharmaceutical composition in an amount sufficient to prevent aninfectious disease in said subject. That is, in some embodiments, thepresent disclosure provides prophylactic vaccines. In some embodiments,the mammalian subject is at risk of exposure to an infectious agent.“Preventing” an infectious disease means to protect a subject fromdeveloping an infectious disease. In some embodiments, preventing aninfectious disease further comprises protecting a subject front beinginfected with an infectious agent (e g., protecting a subject fromdeveloping an acute or a chronic infection). Additionally, the presentdisclosure provides methods of ameliorating a symptom of an infectiousdisease in a mammalian subject in need thereof, comprisingadministration of a pharmaceutical composition in an amount sufficientto ameliorate a symptom of an infectious disease in said subject. Thatis, in some embodiments the present disclosure provides therapeuticvaccines. In some embodiments, the subject is acutely or chronicallyinfected with an infectious agent. The infectious disease may be a viral(e.g., hepatitis, herpes or human papilloma viruses), bacterial, fungal,or parasitic disease. In some embodiments, the pharmaceuticalcomposition further comprises a viral, bacterial, fungal, or parasiticantigen. “Ameliorating” a symptom of an infectious disease means toimprove a symptom preferably diminishing the extent of the disease.

Therapeutic Methods and Uses

The invention further provides therapeutic methods and uses comprisingadministering the compounds of the invention, or pharmaceuticallyacceptable salts thereof, alone or in combination with other therapeuticagents or palliative agents.

In one aspect, the invention provides a method for the treatment ofabnormal cell growth in a subject in need thereof, comprisingadministering to the subject a therapeutically effective amount of acompound of the invention, or a pharmaceutically acceptable saltthereof.

In another aspect, the invention provides a method for the treatment ofabnormal cell growth in a subject in need thereof, comprisingadministering to the subject an amount of a compound of the invention,or a pharmaceutically acceptable salt thereof, in combination with anamount of an additional therapeutic agent (e.g., an anticancertherapeutic agent), which amounts are together effective in treatingsaid abnormal cell growth.

In another aspect, the invention provides a compound of the invention,or a pharmaceutically acceptable salt thereof, for use in the treatmentof abnormal cell growth in a subject.

In a further aspect, the invention provides the use of a compound of theinvention, or a pharmaceutically acceptable salt thereof, for thetreatment of abnormal cell growth in a subject.

In another aspect, the invention provides a pharmaceutical compositionfor use in the treatment of abnormal cell growth in a subject in needthereof, which pharmaceutical composition comprises a compound of theinvention, or a pharmaceutically acceptable salt thereof, and apharmaceutically acceptable carrier or excipient.

In another aspect, the invention provides a compound of the invention,or a pharmaceutically acceptable salt thereof, for use as a medicament,in particular a medicament for the treatment of abnormal cell growth.

In yet another aspect, the invention provides the use of a compound ofthe invention, or a pharmaceutically acceptable salt thereof, for themanufacture of a medicament for the treatment of abnormal cell growth ina subject.

In frequent embodiments of the methods provided herein, the abnormalcell growth is cancer. Compounds of the invention may be administered assingle agents or may be administered in combination with otheranti-cancer therapeutic agents, in particular with standard of careagents appropriate for the particular cancer.

In some embodiments, the methods provided result in one or more of thefollowing effects: (1) inhibiting cancer cell proliferation; (2)inhibiting cancer cell invasiveness; (3) inducing apoptosis of cancercells; (4) inhibiting cancer cell metastasis; or (5) inhibitingangiogenesis.

In some embodiments, the compound of the invention is administered asfirst line therapy. In other embodiments, the compound of the inventionis administered as second (or later) line therapy.

Dosage Forms and Regimens

Administration of the compounds of the invention may be affected by anymethod that enables delivery of the compounds to the site of action.These methods include oral routes, intraduodenal routes, parenteralinjection (including intravenous, subcutaneous, intramuscular,intravascular or infusion), topical, and rectal administration.

Dosage regimens may be adjusted to provide the optimum desired response.For example, a single bolus may be administered, several divided dosesmay be administered overtime or the dose may be proportionally reducedor increased as indicated by the exigencies of the therapeuticsituation. It is especially advantageous to formulate parenteralcompositions in dosage unit form for ease of administration anduniformity of dosage. Dosage unit form, as used herein, refers tophysically discrete units suited as unitary dosages for the mammaliansubjects to be treated; each unit containing a predetermined quantity ofactive compound calculated to produce the desired therapeutic effect inassociation with the required pharmaceutical carrier. The specificationfor the dosage unit forms of the invention are dictated by and directlydependent on (a) the unique characteristics of the chemotherapeuticagent and the particular therapeutic or prophylactic effect to beachieved, and (b) the limitations inherent in the art of compoundingsuch an active compound for the treatment of sensitivity in individuals.

Thus, the skilled artisan would appreciate, based upon the disclosureprovided herein, that the dose and dosing regimen is adjusted inaccordance with methods well known in the therapeutic arts. That is, themaximum tolerable dose can be readily established, and the effectiveamount providing a detectable therapeutic benefit to a patient may alsobe determined, as can the temporal requirements for administering eachagent to provide a detectable therapeutic benefit to the patient.Accordingly, while certain dose and administration regimens areexemplified herein, these examples in no way limit the dose andadministration regimen that may be provided to a patient in practicingthe present invention.

It is to be noted that dosage values may vary with the type and severityof the condition to be alleviated and may include single or multipledoses. It is to be further understood that for any particular subject,specific dosage regimens should be adjusted over time according to theindividual need and the professional judgment of the personadministering or supervising the administration of the compositions, andthat dosage ranges set forth herein are exemplary only and are notintended to limit the scope or practice of the claimed composition. Forexample, doses may be adjusted based on pharmacokinetic orpharmacodynamic parameters, which may include clinical effects such astoxic effects and/or laboratory values. Thus, the present inventionencompasses intra patient dose escalation as determined by the skilledartisan. Determining appropriate dosages and regimens for administrationof the chemotherapeutic agent are well known in the relevant art andwould be understood to be encompassed by the skilled artisan onceprovided the teachings disclosed herein.

The amount of the compound of the invention administered will bedependent on the subject being treated, the severity of the disorder orcondition, the rate of administration, the disposition of the compoundand the discretion of the prescribing physician. However, an effectivedosage is in the range of about 0.001 to about 100 mg per kg body weightper day, preferably about 1 to about 35 mg/kg/day, in single or divideddoses. For a 70 kg human, this would amount to about 0.05 to about 7g/day, preferably about 0.1 to about 2.5 g/day. In some instances,dosage levels below the lower limit of the aforesaid range may be morethan adequate, while in other cases still larger doses may be employedwithout causing any harmful side effect, provided that such larger dosesare first divided into several small doses for administration throughoutthe day.

Formulations and Routes of Administration

As used herein, a “pharmaceutically acceptable carrier” refers to acarrier or diluent that does not cause significant irritation to anorganism and does not abrogate the biological activity and properties ofthe administered compound.

The pharmaceutical acceptable carrier may comprise any conventionalpharmaceutical carrier or excipient. The choice of carrier and/orexcipient will to a large extent depend on factors such as theparticular mode of administration, the effect of the carrier orexcipient on solubility and stability, and the nature of the dosageform.

Suitable pharmaceutical carriers include inert diluents or fillers,water and various organic solvents (such as hydrates and solvates). Thepharmaceutical compositions may, if desired, contain additionalingredients such as flavorings, binders, excipients and the like. Thusfor oral administration, tablets containing various excipients, such ascitric acid may be employed together with various disintegrants such asstarch, alginic acid and certain complex silicates and with bindingagents such as sucrose, gelatin and acacia. Examples, withoutlimitation, of excipients include calcium carbonate, calcium phosphate,various sugars and types of starch, cellulose derivatives, gelatin,vegetable oils and polyethylene glycols. Additionally, lubricatingagents such as magnesium stearate, sodium lauryl sulfate and talc areoften useful for tableting purposes. Solid compositions of a similartype may also be employed in soft and hard filled gelatin capsules.Non-limiting examples of materials, therefore, include lactose or milksugar and high molecular weight polyethylene glycols. When aqueoussuspensions or elixirs are desired for oral administration the activecompound therein may be combined with various sweetening or flavoringagents, coloring matters or dyes and, if desired, emulsifying agents orsuspending agents, together with diluents such as water, ethanol,propylene glycol, glycerin, or combinations thereof.

The pharmaceutical composition may, for example, be in a form suitablefor oral administration as a tablet, capsule, pill, powder, sustainedrelease formulations, solution suspension, for parenteral injection as asterile solution, suspension or emulsion, for topical administration asan ointment or cream or for rectal administration as a suppository.

Exemplary parenteral administration forms include solutions orsuspensions of active compounds in sterile aqueous solutions, forexample, aqueous propylene glycol or dextrose solutions. Such dosageforms may be suitably buffered, if desired.

The pharmaceutical composition may be in unit dosage forms suitable forsingle administration of precise dosages.

Pharmaceutical compositions suitable for the delivery of compounds ofthe invention and methods for their preparation will be readily apparentto those skilled in the art. Such compositions and methods for theirpreparation can be found, for example, in ‘Remington's PharmaceuticalSciences’, 19th Edition (Mack Publishing Company, 1995), the disclosureof which is incorporated herein by reference in its entirety.

The compounds of the invention may be administered orally. Oraladministration may involve swallowing, so that the compound enters thegastrointestinal tract, or buccal or sublingual administration may beemployed by which the compound enters the blood stream directly from themouth.

Formulations suitable for oral administration include solid formulationssuch as tablets, capsules containing particulates, liquids, or powders,lozenges (including liquid filled), chews, multi and nano particulates,gels, solid solution, liposome, films (including muco adhesive), ovules,sprays and liquid formulations.

Liquid formulations include suspensions, solutions, syrups and elixirs.Such formulations may be used as fillers in soft or hard capsules andtypically include a carrier, for example, water, ethanol, polyethyleneglycol, propylene glycol, methylcellulose, or a suitable oil, and one ormore emulsifying agents and/or suspending agents. Liquid formulationsmay also be prepared by the reconstitution of a solid, for example, froma sachet.

The compounds of the invention may also be used in fast dissolving, fastdisintegrating dosage forms such as those described in Expert Opinion inTherapeutic Patents, 11 (6), 981 986 by Liang and Chen (2001), thedisclosure of which is incorporated herein by reference in its entirety.

For tablet dosage forms, depending on dose, the drug may make up from 1wt % to 80 wt % of the dosage form, more typically from 5 wt % to 60 wt% of the dosage form. In addition to the drug, tablets generally containa disintegrant. Examples of disintegrants include sodium starchglycolate, sodium carboxymethyl cellulose, calcium carboxymethylcellulose, croscarmellose sodium, crospovidone, polyvinylpyrrolidone,methyl cellulose, microcrystalline cellulose, lower alkyl substitutedhydroxypropyl cellulose, starch, pregelatinized starch and sodiumalginate. Generally, the disintegrant will comprise from 1 wt % to 25 wt%, preferably from 5 wt % to 20 wt % of the dosage form.

Binders are generally used to impart cohesive qualities to a tabletformulation. Suitable binders include microcrystalline cellulose,gelatin, sugars, polyethylene glycol, natural and synthetic gums,polyvinylpyrrolidone, pregelatinized starch, hydroxypropyl cellulose andhydroxypropyl methylcellulose. Tablets may also contain diluents, suchas lactose (monohydrate, spray dried monohydrate, anhydrous and thelike), mannitol, xylitol, dextrose, sucrose, sorbitol, microcrystallinecellulose, starch and dibasic calcium phosphate dihydrate.

Tablets may also optionally include surface active agents, such assodium lauryl sulfate and polysorbate 80, and glidants such as silicondioxide and talc. When present, surface active agents are typically inamounts of from 0.2 wt % to 5 wt % of the tablet, and glidants typicallyfrom 0.2 wt % to 1 wt % of the tablet.

Tablets also generally contain lubricants such as magnesium stearate,calcium stearate, zinc stearate, sodium stearyl fumarate, and mixturesof magnesium stearate with sodium lauryl sulphate. Lubricants generallyare present in amounts from 0.25 wt % to 10 wt %, preferably from 0.5 wt% to 3 wt % of the tablet.

Other conventional ingredients include anti-oxidants, colorants,flavoring agents, preservatives and taste masking agents.

Exemplary tablets contain up to about 80 wt % drug, from about 10 wt %to about 90 wt % binder, from about 0 wt % to about 85 wt % diluent,from about 2 wt % to about 10 wt % disintegrant, and from about 0.25 wt% to about 10 wt % lubricant.

Tablet blends may be compressed directly or by roller to form tablets.Tablet blends or portions of blends may alternatively be wet, dry, ormelt granulated, melt congealed, or extruded before tableting. The finalformulation may include one or more layers and may be coated oruncoated; or encapsulated.

The formulation of tablets is discussed in detail in “PharmaceuticalDosage Forms: Tablets, Vol. 1”, by H. Lieberman and L. Lachman, MarcelDekker, N.Y., N.Y., 1980 (ISBN 0 8247 6918 X), the disclosure of whichis incorporated herein by reference in its entirety.

Solid formulations for oral administration may be formulated to beimmediate and/or modified release. Modified release formulations includedelayed, sustained pulsed, controlled, targeted and programmed release.

Suitable modified release formulations are described in U.S. Pat. No.6,106,864. Details of other suitable release technologies such as highenergy dispersions and osmotic and coated particles can be found inVerma et al, Pharmaceutical Technology On line, 25(2), 1 14 (2001). Theuse of chewing gum to achieve controlled release is described in WO00/35298. The disclosures of these references are incorporated herein byreference in their entireties.

The compounds of the invention may also be administered directly intothe blood stream, into muscle, or into an internal organ. Suitable meansfor parenteral administration include intravenous, intraarterial,intraperitoneal, intrathecal, intraventricular, intraurethral,intrasternal, intracranial, intramuscular and subcutaneous. Suitabledevices for parenteral administration include needle (including microneedle) injectors, needle free injectors and infusion techniques.

Parenteral formulations are typically aqueous solutions which maycontain excipients such as salts, carbohydrates and buffering agents(preferably to a pH of from 3 to 9), but, for some applications, theymay be more suitably formulated as a sterile non aqueous solution or asa dried form to be used in conjunction with a suitable vehicle such assterile, pyrogen free water.

The preparation of parenteral formulations under sterile conditions, forexample, by lyophilization, may readily be accomplished using standardpharmaceutical techniques well known to those skilled in the art.

The solubility of compounds of the invention used in the preparation ofparenteral solutions may be increased by the use of appropriateformulation techniques, such as the incorporation of solubilityenhancing agents.

Formulations for parenteral administration may be formulated to beimmediate and/or modified release. Modified release formulations includedelayed, sustained, pulsed, controlled, targeted and programmed release.Thus, compounds of the invention may be formulated as a solid, semisolid, or thixotropic liquid for administration as an implanted depotproviding modified release of the active compound. Examples of suchformulations include drug coated stents and PGLA microspheres.

The compounds of the invention may also be administered topically to theskin or mucosa, that is, dermally or transdermally. Typical formulationsfor this purpose include gels, hydrogels, lotions, solutions, creams,ointments, dusting powders, dressings, foams, films, skin patches,wafers, implants, sponges, fibers, bandages and microemulsions.Liposomes may also be used. Typical carriers include alcohol, water,mineral oil, liquid petrolatum, white petrolatum, glycerin, polyethyleneglycol and propylene glycol. Penetration enhancers may be incorporated;see, for example, J Pharm Sci, 88 (10), 955 958 by Finnin and Morgan(October 1999). Other means of topical administration include deliveryby electroporation, iontophoresis, phonophoresis, sonophoresis and microneedle or needle free (e.g. Powderject™, Bioject™, etc.) injection. Thedisclosures of these references are incorporated herein by reference intheir entireties.

Formulations for topical administration may be formulated to beimmediate and/or modified release. Modified release formulations includedelayed, sustained, pulsed, controlled, targeted and programmed release.

The compounds of the invention can also be administered intranasally orby inhalation, typically in the form of a dry powder (either alone, as amixture, for example, in a dry blend with lactose, or as a mixedcomponent particle, for example, mixed with phospholipids, such asphosphatidylcholine) from a dry powder inhaler or as an aerosol sprayfrom a pressurized container, pump, spray, atomizer (preferably anatomizer using electrohydrodynamics to produce a fine mist), ornebulizer, with or without the use of a suitable propellant, such as1,1,1,2 tetrafluoroethane or 1,1,1,2,3,3,3 heptafluoropropane. Forintranasal use, the powder may include a bioadhesive agent, for example,chitosan or cyclodextrin.

The pressurized container, pump, spray, atomizer, or nebulizer containsa solution or suspension of the compound(s) of the invention comprising,for example, ethanol, aqueous ethanol, or a suitable alternative agentfor dispersing, solubilizing, or extending release of the active, apropellant(s) as solvent and an optional surfactant, such as sorbitantrioleate, oleic acid, or an oligolactic acid.

Prior to use in a dry powder or suspension formulation, the drug productis micronized to a size suitable for delivery by inhalation (typicallyless than 5 microns). This may be achieved by any appropriatecomminuting method, such as spiral jet milling, fluid bed jet milling,supercritical fluid processing to form nanoparticles, high pressurehomogenization, or spray drying.

Capsules (made, for example, from gelatin or HPMC), blisters andcartridges for use in an inhaler or insufflator may be formulated tocontain a powder mix of the compound of the invention, a suitable powderbase such as lactose or starch and a performance modifier such as Ileucine, mannitol, or magnesium stearate. The lactose may be anhydrousor in the form of the monohydrate, preferably the latter. Other suitableexcipients include dextran, glucose, maltose, sorbitol, xylitol,fructose, sucrose and trehalose.

A suitable solution formulation for use in an atomizer usingelectrohydrodynamics to produce a fine mist may contain from 1 μg to 20mg of the compound of the invention per actuation and the actuationvolume may vary from 1 μL to 100 μL. A typical formulation includes acompound of the invention, propylene glycol, sterile water, ethanol andsodium chloride. Alternative solvents which may be used instead ofpropylene glycol include glycerol and polyethylene glycol.

Suitable flavors, such as menthol and levomenthol, or sweeteners, suchas saccharin or saccharin sodium, may be added to those formulations ofthe invention intended for inhaled/intranasal administration.

Formulations for inhaled/intranasal administration may be formulated tobe immediate and/or modified release using, for example, poly(DL lacticcoglycolic acid (PGLA). Modified release formulations include delayed,sustained, pulsed, controlled, targeted and programmed release.

In the case of dry powder inhalers and aerosols, the dosage unit isdetermined by means of a valve which delivers a metered amount. Units inaccordance with the invention are typically arranged to administer ametered dose or “puff” containing a desired mount of the compound of theinvention. The overall daily dose may be administered in a single doseor, more usually, as divided doses throughout the day.

Compounds of the invention may be administered rectally or vaginally,for example, in the form of a suppository, pessary, or enema. Cocoabutter is a traditional suppository base, but various alternatives maybe used as appropriate.

Formulations for rectal/vaginal administration may be formulated to beimmediate and/or modified release. Modified release formulations includedelayed, sustained, pulsed, controlled, targeted and programmed release.

Compounds of the invention may also be administered directly to the eyeor ear, typically in the form of drops of a micronized suspension orsolution in isotonic, pH adjusted, sterile saline. Other formulationssuitable for ocular and aural administration include ointments,biodegradable (e.g. absorbable gel sponges, collagen) and nonbiodegradable (e.g. silicone) implants, wafers, lenses and particulateor vesicular systems, such as niosomes or liposomes. A polymer such ascrossed linked polyacrylic acid, polyvinylalcohol, hyaluronic acid, acellulosic polymer, for example, hydroxypropylmethylcellulose,hydroxyethylcellulose, or methyl cellulose, or a heteropolysaccharidepolymer, for example, gelan gum, may be incorporated together with apreservative, such as benzalkonium chloride. Such formulations may alsobe delivered by iontophoresis.

Formulations for ocular/aural administration may be formulated to beimmediate and/or modified release. Modified release formulations includedelayed, sustained, pulsed, controlled, targeted, or programmed release.

Other Technologies

Compounds of the invention may be combined with soluble macromolecularentities, such as cyclodextrin and suitable derivatives thereof orpolyethylene glycol containing polymers, in order to improve theirsolubility, dissolution rate, taste masking, bioavailability and/orstability for use in any of the aforementioned modes of administration.

Drug cyclodextrin complexes, for example, are found to be generallyuseful for most dosage forms and administration routes. Both inclusionand non inclusion complexes may be used. As an alternative to directcomplexation with the drug, the cyclodextrin may be used as an auxiliaryadditive, i.e. as a carrier, diluent, or solubilizer. Most commonly usedfor these purposes are alpha, beta and gamma cyclodextrins, examples ofwhich may be found in PCT Publication Nos. WO 91/11172, WO 94/02518 andWO 98/55148, the disclosures of which are incorporated herein byreference in their entireties.

The amount of the active compound administered will be dependent on thesubject being treated, the severity of the disorder or condition, therate of administration, the disposition of the compound and thediscretion of the prescribing physician. However, an effective dosage istypically in the range of about 0.001 to about 100 mg per kg body weightper day, and frequently about 0.01 to about 35 mg/kg/day, in single ordivided doses. For a 70 kg human, this would amount to about 0.07 mg/dayto about 7000 mg/day, more commonly, from about 10 mg/day to about 1000mg/day. Sometimes, the dosage is about 10, 20, 30, 40, 50, 60, 75, 100,125, 150, 175, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450,475, 500, 525, 550, 575, 600, 625, 650, 675, 700, 750, 800, 900 or 1000mg/day. Sometimes, the dosage is from about 10 mg/day to about 1000mg/day, from about 10 mg/day to about 750 mg/day, from about 10 mg/dayto about 600 mg/day, from about 10 mg/day to about 300 mg/day, fromabout 10 mg/day to about 150 mg/day, from about 20 mg/day to about 750mg/day, from about 20 mg/day to about to 600 mg/day, from about 20mg/day to about to 300 mg/day, from about 20 mg/day to about to 150mg/day, from about 50 mg/day to about 750 mg/day, from about 50 mg/dayto about 600 mg/day, from about 50 mg/day to about 300 mg/day, fromabout 50 mg/day to about 150 mg/day, from about 75 mg/day to about 750mg/day, from about 75 mg/day to about 600 mg/day, from about 75 mg/dayto about 300 mg/day, or from about 75 mg/day to about 150 mg/day.

In some instances, dosage levels below the lower limit of the aforesaidrange may be more than adequate, while in other cases still larger dosesmay be used without causing any harmful side effect, with such largerdoses typically divided into several smaller doses for administrationthroughout the day.

Kit of Parts

Inasmuch as it may desirable to administer a combination of activecompounds, for example, for the purpose of treating a particular diseaseor condition, it is within the scope of the present invention that twoor more pharmaceutical compositions, at least one of which contains acompound in accordance with the invention, may conveniently be combinedin the form of a kit suitable for co-administration of the compositions.Thus, the kit of the invention includes two or more separatepharmaceutical compositions, at least one of which contains a compoundof the invention, and means for separately retaining said compositions,such as a container, divided bottle, or divided foil packet. An exampleof such a kit is the familiar blister pack used for the packaging oftablets, capsules and the like.

The kit of the invention is particularly suitable for administeringdifferent dosage forms, for example, oral and parenteral, foradministering the separate compositions at different dosage intervals,or for titrating the separate compositions against one another. Toassist compliance, the kit typically includes directions foradministration and may be provided with a memory aid.

Combination Therapy

As used herein, the term “combination therapy” refers to theadministration of a compound of the invention together with an at leastone additional pharmaceutical or medicinal agent (e.g., an anti-cancertherapeutic agent), either sequentially or simultaneously.

As noted herein, the compounds of the invention may be used incombination with one or more additional anti-cancer therapeutic agent.The efficacy of the compounds of the invention in certain tumors may beenhanced by combination with other approved or experimental cancertherapies, e.g., radiation, surgery, chemotherapeutic agents, targetedtherapies, agents that inhibit other signaling pathways that aredysregulated in tumors, and other immune enhancing agents, such as PD 1or PD L1 antagonists and the like.

When a combination therapy is used, the one or more additionalanti-cancer therapeutic agents may be administered sequentially orsimultaneously with the compound of the invention. In one embodiment,the additional anti-cancer therapeutic agent is administered to a mammal(e.g., a human) prior to administration of the compound of theinvention. In another embodiment, the additional anti-cancer therapeuticagent is administered to the mammal after administration of the compoundof the invention. In another embodiment, the additional anti-cancertherapeutic agent is administered to the mammal (e.g., a human)simultaneously with the administration of the compound of the invention.

The invention also relates to a pharmaceutical composition for thetreatment of abnormal cell growth in a mammal, including a human, whichcomprises an amount of a compound of the invention, as defined above(including hydrates, solvates and polymorphs of said compound orpharmaceutically acceptable salts thereof), in combination with one ormore (preferably one to three) additional anti-cancer therapeuticagents.

Classes of additional chemotherapeutic agents, which can be administeredin combination with a compound of this invention, include, but are notlimited to: alkylating agents, antimetabolites, kinase inhibitors,spindle poison plant alkaloids, cytotoxic/antitumor antibiotics,topisomerase inhibitors, photosensitizers, anti-estrogens and selectiveestrogen receptor modulators (SERMs), anti-progesterones, estrogenreceptor down-regulators (ERDs), estrogen receptor antagonists,leutinizing hormone-releasing hormone agonists; IL-2 receptor agonist(recombinant cytokines or agonists for cytokine receptors); andanti-sense oligonucleotides or oligonucleotides derivatives that inhibitexpression of genes implicated in abnormal cell proliferation or tumorgrowth.

Other additional chemotherapy agents include not only taxanes orplatinum agents but also HER2 targeted agents, e.g., trastuzumab.

In another embodiment, such additional anti-cancer therapeutic agentsinclude compounds derived from the following classes: mitoticinhibitors, alkylating agents, antimetabolites, antitumor antibiotics,anti-angiogenesis agents, topoisomerase I and II inhibitors, plantalkaloids, spindle poison plant alkaloids, KRAS inhibitors; MCT4inhibitors; MAT2a inhibitors; alk/c-Met/ROS inhibitors (includingcrizotinib or lorlatinib); mTOR inhibitors (including temsirolimus orgedatolisib); src/abl inhibitors (including bosutinib); cyclin-dependentkinase (CDK) inhibitors (including palbociclib, PF-06873600); erbinhibitors (including dacomitinib); PARP inhibitors (includingtalazoparib); SMO inhibitors (including glasdegib); EGFR T790Minhibitors; PRMT5 inhibitors; TGFβR1 inhibitors; growth factorinhibitors; cell cycle inhibitors, biological response modifiers; enzymeinhibitors; and cytotoxics.

In another embodiment, such additional anti-cancer therapeutic agentsinclude compounds derived from an anti-angiogenesis agent, including forexample tyrosine kinase/vascular endothelial growth factor (VEGF)receptor (VEGFR) inhibitors (including sunitinib, axitinib, sorafenib,and tivozanib), TIE-2 inhibitors, PDGFR inhibitors, angiopoetininhibitors, PKCp inhibitors, COX-2 (cyclooxygenase II) inhibitors,integrins (alpha-v/beta-3), MMP-2 (matrix-metalloproteinase 2)inhibitors, and MMP-9 (matrix-metalloproteinase 9) inhibitors. Preferredanti-angiogenesis agents include sunitinib (Sutent™), bevacizumab(Avastin™), axitinib (Inlyta™) SU 14813 (Pfizer), and AG 13958 (Pfizer).Additional anti-angiogenesis agents include vatalanib (CGP 79787),pegaptanib octasodium (Macugen™), vandetanib (Zactima™), PF-0337210(Pfizer), SU 14843 (Pfizer), AZD 2171 (AstraZeneca), ranibizumab(Lucentis™), Neovastat™ (AE 941), tetrathiomolybdata (Coprexa™), AMG 706(Amgen), VEGF Trap (AVE 0005), CEP 7055 (Sanofi-Aventis), XL 880(Exelixis), telatinib (BAY 57-9352), and CP-868,596 (Pfizer). Otheranti-angiogenesis agents include enzastaurin (LY 317615), midostaurin(CGP 41251), perifosine (KRX 0401), teprenone (Selbex™) and UCN 01(Kyowa Hakko). Other examples of anti-angiogenesis agents includecelecoxib (Celebrex™), parecoxib (Dynastat™), deracoxib (SC 59046),lumiracoxib (Preige™), valdecoxib (Bextra™), rofecoxib (Vioxx™),iguratimod (Careram™), IP 751 (Invedus), SC-58125 (Pharmacia) andetoricoxib (Arcoxia™). Yet further anti-angiogenesis agents includeexisulind (Aptosyn™), salsalate (Amigesic™), diflunisal (Dolobid™),ibuprofen (Motrin™), ketoprofen (Orudis™), nabumetone (Relafen™),piroxicam (Feldene™), naproxen (Aleve™, Naprosyn™), diclofenac(Voltaren™), indomethacin (Indocin™) sulindac (Clinoril™), tolmetin(Tolectin™), etodolac (Lodine™), ketorolac (Toradol™), and oxaprozin(Daypro™). Yet further anti-angiogenesis agents include ABT 510(Abbott), apratastat (TMI 005), AZD 8955 (AstraZeneca), incyclinide(Metastat™), and PCK 3145 (Procyon). Yet further anti-angiogenesisagents include acitretin (Neotigason™), plitidepsin (Aplidine™)cilengtide (EMD 121974), combretastatin A4 (CA4P), fenretinide (4 HPR),halofuginone (Tempostatin™), Panzem™ (2-methoxyestradiol), PF-03446962(Pfizer), rebimastat (BMS 275291), catumaxomab (Removab™), lenalidomide(Revlimid™), squalamine (EVIZON™) thalidomide (Thalomid™), Ukrain™ (NSC631570), Vitaxin™ (MEDI 522), and zoledronic acid (Zometa™).

In another embodiment, such additional anti-cancer therapeutic agentsinclude compounds derived from hormonal agents and antagonists. Examplesinclude where anti-hormonal agents act to regulate or inhibit hormoneaction on tumors such as anti-estrogens and selective estrogen receptormodulators (SERMs), and a selective estrogen receptor degrader (SERD)including tamoxifen, raloxifene, droloxifene, 4-hydroxytamoxifen,trioxifene, keoxifene, LY117018, onapristone, toremifene (Fareston), andfulvestrant. Examples also include aromatase inhibitors that inhibit theenzyme aromatase, which regulates estrogen production in the adrenalglands, and include compounds like 4(5)-imidazoles, aminoglutethimide,megestrol acetate, exemestane, formestane, fadrozole, vorozole,letrozole, and anastrozole; and anti-androgens such as flutamide,nilutamide, bicalutamide, leuprolide, fluridil, apalutamide,enzalutamide, cimetidine and goserelin.

In another embodiment, such additional anti-cancer therapeutic agentsinclude compounds derived from signal transduction inhibitors, such asinhibitors of protein tyrosine kinases and/or serine/threonine kinases:a signal transduction inhibitor (e.g., inhibiting the means by whichregulatory molecules that govern the fundamental processes of cellgrowth, differentiation, and survival communicated within the cell).Signal transduction inhibitors include small molecules, antibodies, andantisense molecules. Signal transduction inhibitors include for examplekinase inhibitors (e.g., tyrosine kinase inhibitors or serine/threoninekinase inhibitors) and cell cycle inhibitors. More specifically signaltransduction inhibitors include, for example, farnesyl proteintransferase inhibitors, EGF inhibitor, ErbB-1 (EGFR), ErbB-2, pan erb,IGF1R inhibitors, MEK (including binimetinib (Mektovi™)), c-Kitinhibitors, FLT-3 inhibitors, K-Ras inhibitors, PI3 kinase inhibitors,JAK inhibitors, STAT inhibitors, Raf kinase inhibitors, BRAF (includingencorafenib (Braftovi™)), Akt inhibitors, mTOR inhibitor, P70S6 kinaseinhibitors, inhibitors of the WNT pathway and multi-targeted kinaseinhibitors.

In another embodiment, such additional anti-cancer therapeutic agentsinclude docetaxel, paclitaxel, paclitaxel protein-bound particles,cisplatin, carboplatin, oxaliplatin, capecitabine, gemcitabine orvinorelbine.

In another embodiment, such additional anti-cancer therapeutic agentsinclude compounds derived from an epigenetic modulator, where examplesinclude an inhibitor of EZH2 (including PF-06821497), SMARCA4, PBRM1,ARID1A, ARID2, ARID1B, DNMT3A, TET2, MLL1/2/3, NSD1/2, SETD2, BRD4,DOT1L, HKMTsanti, PRMT1-9, LSD1, UTX, IDH1/2 or BCL6.

In another embodiment, such additional anti-cancer therapeutic agentsinclude compounds that are immuno-oncology agents, includingimmunomodulatory agents.

In another embodiment, combinations with pattern recognition receptors(PRRs) are contemplated. PRRs are receptors that are expressed by cellsof the immune system and that recognize a variety of moleculesassociated with pathogens and/or cell damage or death. PRRs are involvedin both the innate immune response and the adaptive immune response. PRRagonists may be used to stimulate the immune response in a subject.There are multiple classes of PRR molecules, including toll-likereceptors (TLRs), RIG-1-like receptors (RLRs), nucleotide-bindingoligomerization domain (NOD)-like receptors (NLRs), C-type lectinreceptors (CLRs), and Stimulator of Interferon Genes (STING) protein.

The STING protein functions as both a cytosolic DNA sensor and anadaptor protein in Type 1 interferon signaling. The terms “STING” and“stimulator of interferon genes” refer to any form of the STING protein,as well as variants, isoforms, and species homologs that retain at leasta part of the activity of STING. Unless indicated differently, such asby specific reference to human STING, STING includes all mammalianspecies of native sequence STING, e.g. human, monkey, and mouse STING isalso known as—TMEM173.

“STING agonist” as used herein means, any molecule, which upon bindingto STING, (1) stimulates or activates STING, (2) enhances, increases,promotes, induces, or prolongs an activity, function, or presence ofSTING, or (3) enhances, increases, promotes, or induces the expressionof STING. STING agonists useful in the any of the treatment method,medicaments and uses of the present invention include, for example,nucleic acid ligands which bind STING.

Examples of STING agonists that are useful in the treatment methods,medicaments, and uses of the present invention include variousimmunostimulatory nucleic acids, such as synthetic double stranded DNA,cyclic di-GMP, cyclic-GMP-AMP (cGAMP), synthetic cyclic dinucleotides(CDN) such as MK-1454 and ADU-S100 (MIW815), and small molecules such asWO2019027858, WO20180093964, WO2017175156, WO2017175147.

Therapeutic antibodies may have specificity against a variety ofdifferent antigens. For example, therapeutic antibodies may be directedto a tumor associated-antigen, such that binding of the antibody to theantigen promotes death of the cell expressing the antigen. In otherexample, therapeutic antibodies may be directed to an antigen on animmune cell, such that binding of the antibody prevents downregulationof the activity of the cell expressing the antigen (and thereby promotesactivity of the cell expressing the antigen). In some situations, atherapeutic antibody may function through multiple different mechanisms(for example, it may both i) promote death of the cell expressing theantigen, and ii) prevent the antigen from causing down-regulation of theactivity of immune cells in contact with the cell expressing theantigen).

In another embodiment, such additional anti-cancer therapeutic agentsinclude antibodies that would be blocking or inhibitory at the target:CTLA-4 (including ipilimumab or tremelimumab), PD-1 or PD-L1 (includingatezolizumab, avelumab, cemiplimab, durvalumab, nivolumab, orpembrolizumab), LAG-3, TIM-3, or TIGIT.

In another embodiment, such additional anti-cancer therapeutic agentsinclude antibodies that are agonists of 4-1BB, OX40, GITR, ICOS, orCD40.

In another embodiment the anti-cancer therapy may be a CAR-T-celltherapy.

Examples of a therapeutic antibody include: an anti-OX40 antibody, ananti-4-1BB antibody, an anti-HER2 antibody (including an anti-HER2antibody-drug conjugate (ADC)), a bispecific anti-CD47/anti-PD-L1antibody, and a bispecific anti-P-cadherin/anti-CD3 antibody. Examplesof cytotoxic agents that may be incorporated in an ADC include ananthracycline, an auristatin, a dolastatin, a combretastatin, aduocarmycin, a pyrrolobenzodiazepine dimer, an indolino-benzodiazepinedimer, an enediyne, a geldanamycin, a maytansine, a puromycin, a taxane,a vinca alkaloid, a camptothecin, a tubulysin, a hemiasterlin, aspliceostatin, a pladienolide, and stereoisomers, isosteres, analogs, orderivatives thereof. Exemplary immunomodulating agents that may beincorporated in an ADC include gancyclovier, etanercept, tacrolimus,sirolimus, voclosporin, cyclosporine, rapamycin, cyclophosphamide,azathioprine, mycophenolgate mofetil, methotrextrate, glucocorticoid andits analogs, cytokines, stem cell growth factors, lymphotoxins, tumornecrosis factor (TNF), hematopoietic factors, interleukins (e.g.,interleukin-1 (IL-1), IL-2, IL-3, IL-6, IL-10, IL-12, IL-15, IL-18, andIL-21), colony stimulating factors (e.g., granulocyte-colony stimulatingfactor (G-CSF) and granulocyte macrophage-colony stimulating factor(GM-CSF)), interferons (e.g., interferons-.alpha., -.beta. and -.gamma),the stem cell growth factor designated “S 1 factor,” erythropoietin andthrombopoietin, or a combination thereof.

Additional examples of therapeutic antibodies may include the followingantigens where exemplary antibodies directed to the antigen are alsoincluded below (in brackets/parenthesis after the antigen). The antigensas follow may also be referred to as “target antigens” or the likeherein. Target antigens for therapeutic antibodies herein include, forexample: 4-1BB (e.g. utomilumab); 5T4; A33; alpha-folate receptor 1(e.g. mirvetuximab soravtansine); Alk-1; BCMA [e.g. see U.S. Pat. No.9,969,809]; BTN1A1 (e.g. see WO2018222689); CA-125 (e.g. abagovomab);Carboanhydrase IX; CCR2; CCR4 (e.g. mogamulizumab); CCR5 (e.g.Ieronlimab); CCR8; CD3 [e.g. blinatumomab (CD3/CD19 bispecific),CD3/P-cadherin bispecific, CD3/BCMA bispecific] CD19 (e.g. blinatumomab,MOR208); CD20 (e.g. ibritumomab tiuxetan, obinutuzumab, ofatumumab,rituximab, ublituximab); CD22 (inotuzumab ozogamicin, moxetumomabpasudotox); CD25; CD28; CD30 (e.g. brentuximab vedotin); CD33 (e.g.gemtuzumab ozogamicin); CD38 (e.g. daratumumab, isatuximab), CD40;CD-40L; CD44v6; CD47 (e.g. Hu5F9-G4, CC-90002, SRF231, B6H12); CD52(e.g. alemtuzumab); CD56; CD63; CD79 (e.g. polatuzumab vedotin); CD80;CD123; CD276/B7-H3 (e.g. omburtamab); CDH17; CEA; CIhCG; CTLA-4 (e.g.ipilimumab, tremelimumab), CXCR4; desmoglein 4; DLL3 (e.g.rovalpituzumab tesirine); DLL4; E-cadherin; EDA; EDB; EFNA4; EGFR (e.g.cetuximab, depatuxizumab mafodotin, necitumumab, panitumumab); EGFRvlll;Endosialin; EpCAM (e.g. oportuzumab monatox); FAP; Fetal AcetylcholineReceptor; FLT3 (e.g. see WO2018/220584); GD2 (e.g. dinutuximab, 3F8);GD3; GITR; GloboH; GM1; GM2; HER2/neu [e.g. margetuximab, pertuzumab,trastuzumab; ado-trastuzumab emtansine, trastuzumab duocarmazine, [seeU.S. Pat. No. 8,828,401]; HER3; HER4; ICOS; IL-10; ITG-AvB6; LAG-3 (e.g.relatlimab); Lewis-Y; LG; Ly-6; M-CSF [see U.S. Pat. No. 7,326,414];MCSP; mesothelin; MUC1; MUC2; MUC3; MUC4; MUCSAC; MUC5B; MUC7; MUC16;Notch1; Notch3; Nectin-4 (e.g. enfortumab vedotin); OX40 [see U.S. Pat.No. 7,960,515]; P-Cadherein [see WO2016/001810]; PCDHB2; PDGFRA (e.g.olaratumab); Plasma Cell Antigen; PolySA; PSCA; PSMA; PTK7 [see U.S.Pat. No. 9,409,995]; Ror1; SAS; SCRx6; SLAMF7 (e.g. elotuzumab); SHH;SIRPa (e.g. ED9, Effi-DEM); STEAP; TGF-beta; TIGIT; TIM-3; TMPRSS3;TNF-alpha precursor; TROP-2 (e.g sacituzumab govitecan); TSPAN8; VEGF(e.g. bevacizumab, brolucizumab); VEGFR1 (e.g. ranibizumab); VEGFR2(e.g. ramucirumab, ranibizumab); Wue-1.

Exemplary imaging agents that may be included in an ADC includefluorescein, rhodamine, lanthanide phosphors, and their derivativesthereof, or a radioisotope bound to a chelator. Examples of fluorophoresinclude, but are not limited to, fluorescein isothiocyanate (FITC)(e.g., 5-FITC), fluorescein amidite (FAM) (e.g., 5-FAM), eosin,carboxyfluorescein, erythrosine, Alexa Fluor@ (e.g., Alexa 350, 405,430, 488, 500, 514, 532, 546, 555, 568, 594, 610, 633, 647, 660, 680,700, or 750), carboxytetramethylrhodamine (TAMRA) (e.g., 5-TAMRA),tetramethylrhodamine (TMR), and sulforhodamine (SR) (e.g., SR101).Examples of chelators include, but are not limited to,1,4,7,10-tetraazacyclododecane-N,N′,N″,N′″-tetraacetic acid (DOTA),1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA),1,4,7-triazacyclononane, 1-glutaric acid-4,7-acetic acid (deferoxamine),diethylenetriaminepentaacetic acid (DTPA), and1,2-bis(o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid) (BAPTA).

Exemplary therapeutic proteins that may be included in an ADC include atoxin, a hormone, an enzyme, and a growth factor.

Exemplary biocompatible polymers that may be incorporated in an ADCinclude water-soluble polymers, such as polyethylene glycol (PEG) or itsderivatives thereof and zwitterion-containing biocompatible polymers(e.g., a phosphorylcholine containing polymer).

Exemplary biocompatible polymers that may be incorporated in an ADCinclude anti-sense oligonucleotides.

The invention also concerns the use of radiation in combination with anyanti-cancer therapeutic agent administered herein. More specifically,compounds of the invention can be administered in combination withadditional therapies, such as radiation therapy and/or chemotherapy.

Chemical Synthesis

The following schemes and written descriptions provide general detailsregarding the preparation of the compounds of the invention.

The compounds of the invention may be prepared by any method known inthe art for the preparation of compounds of analogous structure. Inparticular, the compounds of the invention can be prepared by theprocedures described by reference to the Schemes that follow, or by thespecific methods described in the Examples, or by similar processes toeither.

The skilled person will appreciate that the experimental conditions setforth in the schemes that follow are illustrative of suitable conditionsfor effecting the transformations shown, and that it may be necessary ordesirable to vary the precise conditions employed for the preparation ofcompounds of Formula (I), and compounds that fall within Formula (I),e.g., compounds of Formulas (I), (Ia) or (Ib), and the like.

In addition, the skilled person will appreciate that it may be necessaryor desirable at any stage in the synthesis of compounds of the inventionto protect one or more sensitive groups, to prevent undesirable sidereactions. In particular, it may be necessary or desirable to protectamino or alcohol groups. The protecting groups (PGs) used in thepreparation of the compounds of the invention may be used inconventional manner. See, for example, those described in ‘Greene'sProtective Groups in Organic Synthesis’ by Theodora W Greene and Peter GM Wuts, third edition, (John Wiley and Sons, 1999), in particularchapters 7 (“Protection for the Amino Group”) and 2 (“Protection for theHydroxyl Group, Including 1,2- and 1,3-Diols”), incorporated herein byreference, which also describes methods for the removal of such groups.

All of the derivatives of Formula (I) can be prepared by the proceduresdescribed in the general methods presented below or by routinemodifications thereof. The present invention also encompasses any one ormore of these processes for preparing the derivatives of Formula (I), inaddition to any novel intermediates used therein. The person skilled inthe art will appreciate that the following reactions may be heatedthermally or under microwave irradiation or under flow chemistryconditions.

It will be further appreciated that it may be necessary or desirable tocarry out the transformations in a different order from that describedin the schemes, or to modify one or more of the transformations, toprovide the desired compound of the invention.

According to a first process, compounds of Formula (I) may be preparedfrom compounds of intermediate (i) as illustrated by Scheme 1.

Possible PGs include 4-methoxybenzyl, N,N-bis-(4-methoxybenzyl),tert-octyl or other suitable amine protecting group;N,N-bis-(4-methoxybenzyl) has been used most frequently for compounds ofFormula (I).

Intermediates (i), (iv), (vi), (ix), (x) are commercially available ormay be synthesized by those of ordinary skill in the art according tothe literature or preparations described herein. For the Schemesdiscussed herein, when compounds of Formula (I) have chiral centers, therespective enantiomers may be separated via chiral separation of theracemate as required. Also, when R³ contains a protecting group such asa ketal or silyl, suitable deprotection conditions may be employed asnecessary, such as methanesulfonic acid indichloromethane/methanol/water.

Intermediate (ii) may be prepared from intermediate (i) according tostep (a), a nitration reaction. Typical methods employ use of a suitablenitrating agent and suitable organic or inorganic solvent. Preferredconditions comprise use of nitric acid in sulfuric acid at 10° C.

Intermediate (iii) may be prepared from intermediate (ii) according tostep (b), a triflate forming reaction. Typical methods employ use oftrifluoromethanesulfonic anhydride with a suitable organic or inorganicbase in a suitable organic solvent at reduced temperatures. Preferredconditions comprise use of triethylamine in dichloroethane at 0° C.

Intermediate (v) may be prepared from intermediate (iii) according tostep (c), a nucleophilic aromatic substitution reaction withintermediate (iv). Typical methods employ use of a suitable organic orinorganic base in a suitable organic solvent at RT or elevatedtemperatures either thermally, under microwave irradiation, or flowchemistry conditions. Preferred conditions comprise use of triethylaminein dichloroethane.

Intermediate (vii) may be prepared from intermediate (v) according tostep (d), a nucleophilic aromatic substitution reaction withintermediate (vi). Typical methods employ use of a suitable organic orinorganic base in a suitable organic solvent at RT or elevatedtemperatures either thermally, under microwave irradiation, or flowchemistry conditions. For example, use of Bis(4-methoxybenzyl)amine withtriethylamine in dichloroethane at 50° C. heated thermally or use oftert-Octylamine with triethylamine in toluene at 75° C.

Intermediate (viii) may be prepared from intermediate (vii) according tostep (e), a nitro reduction step. Typical methods employ hydrogenationconditions with a suitable hydrogen source and suitable hydrogenationcatalyst in a suitable organic solvent at RT or at elevated temperaturesheated thermally, under microwave irradiation, or flow chemistryconditions or use of a suitable metal and suitable hydrogen or protondonor in a suitable organic solvent. Preferred conditions comprise useof zinc dust and ammonium formate in methanol.

Intermediate (xi) may be prepared from intermediate (viii) according tostep (f), an amide bond forming step with intermediate (ix) or (x).Typical methods employ use of intermediate (ix) with a suitable organicor inorganic base in a suitable organic solvent or use of intermediate(x) with a suitable amide coupling reagent and a suitable organic orinorganic base in a suitable organic solvent. Typical conditions usetriethylamine and dichloromethane when using intermediate (ix). Typicalconditions use 50 wt % propylphosphonic acid anhydride solution inethylacetate, triethylamine and ethyl acetate when using intermediate(x).

Intermediate (xii) may be prepared from intermediate (xi) according tostep (g), an imidazole ring formation. Typical methods employ basicconditions utilizing a suitable organic or inorganic base at elevatedtemperatures either thermally or under microwave irradiation or flowchemistry conditions; alternatively, acidic conditions utilizing asuitable organic or inorganic acid at elevated temperatures eitherthermally or under microwave irradiation or flow chemistry conditions.Alternatively, a suitable dehydrating agent in a suitable organicsolvent at elevated temperatures either thermally or under microwaveirradiation or flow chemistry conditions. Preferred conditions compriseuse of sodium hydroxide in ethanol at 80° C. heated thermally ortriphenylphosphine and triethylamine in carbon tetrachloride at 80° C.heated thermally.

Compounds of Formula (I) may be prepared from intermediate (xii)according to step (h), a removal of PG and those contained in R³ ifpresent, eg. silyl or ketal. Typical deprotection methods comprise asuitable organic or inorganic acid in a suitable organic solvent at RTor at elevated temperatures either thermally or under microwaveirradiation or flow chemistry conditions. Preferred conditions comprisemethanesulfonic acid in dichloromethane at 45° C. heated thermallyfollowed by addition of methanol and water.

Alternatively, compounds of Formula (I) may be prepared fromintermediate (iii), as illustrated by Scheme 2.

Possible amino PGs include 4-methoxybenzyl, N,N-bis(4-methoxybenzyl),tert-octyl, with tert-octyl being used most frequently in the examples.

A leaving group (LG) is a functional group to assist with a specificreaction and includes OH, Cl, Br, I, OMs, OTs, and OTf, with OH beingused most frequently in the examples.

Intermediates (ix), (x), (xiii), (xv) are commercially available or maybe synthesized by those skilled in the art according to the literatureor preparations described herein.

Intermediate (xiv) may be prepared from intermediate (iii) according tostep (i), a nucleophilic aromatic substitution reaction withintermediate (xiii). Typical methods employ use of a suitable organic orinorganic base in a suitable organic solvent at RT or elevatedtemperatures either thermally, under microwave irradiation, or flowchemistry conditions. Preferred conditions comprise use of triethylaminein dichloroethane.

Intermediate (xvi) may be prepared from intermediate (xiv) according tostep (j), a nucleophilic aromatic substitution reaction withintermediate (xv). Typical methods employ use of a suitable organic orinorganic base in a suitable organic solvent at RT or elevatedtemperatures either thermally, under microwave irradiation, or flowchemistry conditions. Preferred conditions comprise use ofbis(4-methoxybenzyl)amine with triethylamine in dichloroethane at 50° C.heated thermally or use of tert-Octylamine with triethylamine in tolueneat 75° C.

Intermediate (xvii) may be prepared from intermediate (xvi) according tostep (k), a tandem nitro reduction and debenzylation step. Typicalmethods employ hydrogenation conditions with a suitable hydrogen sourceand suitable hydrogenation catalyst in a suitable organic solvent at RTor at elevated temperatures heated thermally, under microwaveirradiation, or flow chemistry conditions. Preferred conditions compriseuse of ammonium formate and 30% palladium on carbon in ethanol at 55° C.

Intermediate (xviii) may be prepared from intermediate (xvii) accordingto step (I), an amide bond forming step with intermediate (ix) or (x).Typical methods employ use of intermediate (ix) with a suitable organicor inorganic base in a suitable organic solvent or use of intermediate(x) with a suitable amide coupling reagent and a suitable organic orinorganic base in a suitable organic solvent. Preferred conditionscomprise use of intermediate (ix) with triethylamine and dichloromethaneat 0° C.

Intermediate (xix) may be prepared from intermediate (xviii) accordingto step (m), an imidazole ring formation. Typical methods employ basicconditions utilizing a suitable organic or inorganic base at elevatedtemperatures either thermally, under microwave irradiation, or flowchemistry conditions; acidic conditions utilizing a suitable organic orinorganic acid at elevated temperatures either thermally, undermicrowave irradiation, or flow chemistry conditions, or a suitabledehydrating agent in a suitable organic solvent at elevated temperatureseither thermally, under microwave irradiation, or flow chemistryconditions. Preferred conditions comprise use of sodium hydroxide inethanol at 75° C. heated thermally

Intermediate (xxi) may be prepared from intermediate (xix) according tostep (n), a nucleophilic substitution reaction or Mitsunobu reactionwith intermediate (xx). Typical methods comprise a suitable organic orinorganic base in a suitable organic solvent at RT or elevatedtemperatures either thermally, under microwave irradiation, or flowchemistry conditions. Alternatively, when LG is a hydroxyl group bytreatment with a suitable phosphine and suitable azodicarboxylate (orcombination of both in a single reagent) in a suitable organic solventat RT or elevated temperatures either thermally, under microwaveirradiation, or flow chemistry conditions. Preferred conditions, whereLG is a hydroxyl group, comprise use ofcyanomethylenetributylphosphorane in toluene at 90° C. or 100° C. heatedthermally.

Compounds of Formula (I) may be prepared from intermediate (xxi)according to step (o), a removal of PG and any PG contained in R³ ifpresent, for example a ketal or silane. Typical methods comprise asuitable organic or inorganic acid in a suitable organic solvent at RTor at elevated temperatures either thermally, under microwaveirradiation, or flow chemistry conditions. Preferred conditions comprisemethanesulfonic acid in hexafluoroisopropanol or trifluoroacetic acid indichloromethane followed by addition of methanol.

In executing the synthesis of the compounds of the invention, oneskilled in the art will monitor reactions with common methods thatinclude thin-layer chromatography (TLC), liquid chromatography/massspectroscopy (LCMS), and nuclear magnetic resonance (NMR).

One skilled in the art will also recognize that the compounds of theinvention may be prepared as mixtures of diastereomers or geometricisomers (e.g., cis and trans substitution on a cycloalkane ring). Theseisomers can be separated by standard chromatographic techniques, such asnormal phase chromatography on silica gel, reverse phase preparativehigh pressure liquid chromatography or supercritical fluidchromatography. One skilled in the art will also recognize that somecompounds of the invention are chiral and thus may be prepared asracemic or scalemic mixtures of enantiomers. Several methods areavailable and are well known to those skilled in the art for theseparation of enantiomers.

EXAMPLES

Except where otherwise noted, reactions were run under an atmosphere ofnitrogen. Chromatography on silica gel was carried out using 250-400mesh silica gel using pressurized nitrogen (˜10-15 psi) to drive solventthrough the column (“flash chromatography”). Where indicated, solutionsand reaction mixtures were concentrated by rotary evaporation undervacuum.

¹H and ¹⁹F Nuclear magnetic resonance (NMR) spectra were in all casesconsistent with the proposed structures. Characteristic chemical shifts(5) are given in parts-per-million downfield from tetramethylsilane (for¹H-NMR) using conventional abbreviations for designation of major peaks:e.g. s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet; br,broad. The following abbreviations have been used for common solvents:CDCl₃, deuterochloroform; d₆-DMSO, deuterodimethylsulfoxide; and CD₃OD,deuteromethanol. Where appropriate, tautomers may be recorded within theNMR data; and some exchangeable protons may not be visible.

Mass spectra, MS (m/z), were recorded using either electrosprayionization (ESI) or atmospheric pressure chemical ionization (APCI).Where relevant and unless otherwise stated, the m/z data provided arefor isotopes ¹⁹F, ³⁵Cl, ⁷⁹Br and ¹²⁷I.

The nomenclature is written as described by IUPAC (International Unionof Pure and Applied Chemistry generated within Perkin Elmers Chemdraw18.0.

In the non-limiting Examples and Preparations that are set out herein,the following the abbreviations apply:

AcOH is acetic acid;

aq is aqueous;

Bn is benzyl;

br is broad;

tBu is tert-butyl;

° C. is degrees Celsius;

CO₂ is carbon dioxide;

CMBP is Cyanomethylenetributylphosphorane;

Cs₂CO₃ is cesium carbonate;

DCE is dichloroethane;

DCM is dichloromethane; methylene chloride;

DIPEA/DIEA is N-ethyldiisopropylamine, N,N-diisopropylethylamine;

DMA is dimethylacetamide;

DMF is N,N-dimethylformamide;

DMSO is dimethyl sulphoxide;

ee is enantiomeric excess;

EtOAc is ethyl acetate;

EtOH is ethanol;

Et₃N is triethylamine;

g is gram;

HCO₂NH₄ is ammonium formate;

HCl is hydrochloric acid;

HFIP is 1,1,1,3,3,3-hexafluoroisopropanol;

HNO₃ is nitric acid;

HPLC is high pressure liquid chromatography;

H₂O is water;

H₂SO₄ is sulfuric acid;

Hr or hr is hour;

IPA/iPrOH is isopropanol;

L is litre;

LCMS is liquid chromatography mass spectrometry;

LiAlH₄ is lithium aluminium hydride;

LiOH is lithium hydroxide;

M is molar;

MeCN is acetonitrile;

MeI is methyl iodide;

MeOH is methanol;

mg is milligram;

MgSO₄ is magnesium sulphate;

MHz is mega Hertz;

min is minutes;

mL is milli litre;

mmol is millimole;

mol is mole;

MS m/z is mass spectrum peak;

MsOH is methanesulfonic acid;

NaH is sodium hydride;

NaHCO₃ is sodium hydrogencarbonate;

NaOH is sodium hydroxide;

Na₂SO₄ is sodium sulphate;

NH₃ is ammonia;

NH₄OH is ammonium hydroxide;

NH(PMB)₂ is bis(4-methoxybenzyl)amine;

NMR is nuclear magnetic resonance;

Pd/C is palladium on carbon;

pH is power of hydrogen;

ppm is parts per million;

psi is pounds per square inch;

Rt is retention time;

RT is room temperature;

TBDMS is tertbutyldimethylsilyl;

TBSCI is tertbutylimethylsilyl chloride;

TBME/MTBE is tert-butyl dimethyl ether;

TEA is triethylamine;

Tf₂O is trifluoromethanesulfonic anhydride;

TFA is trifluoroacetic acid;

TFAA is trifluoroacetic anhydride;

THF is tetrahydrofuran;

TLC is thin layer chromatography;

TsOH is p-Toluenesulfonic acid;

Zn is zinc;

μL is microlitre;

μmol is micromol

Chiral separations were used to separate enantiomers of someintermediates during the preparation of the compounds of the invention.When such separation was done, the separated enantiomers were designatedas ENT-1 or ENT-2, according to their order of elution. For compoundswith two chiral centers, the stereoisomers at each stereocenter wereseparated at different times. The designation of ENT-1 or ENT-2 of anintermediate or an example refers to the chiral center for theseparation done at that step. It is recognized that when stereoisomersat a chiral center are separated in a compound with two or more centers,the separated enantiomers are diastereomers of each other. The ENT-1 orENT-2 designation is used herein for consistency and refers to theseparated chiral center. By way of example, but not limitation, Examples6 and 7 have a chiral center. The enantiomers were separated as the laststep. The chiral center is drawn as the two possibilities, but it is notknown which example is which enantiomer. Therefore, the (R) and (S)designation is not associated with either Example 6 or 7. If theseparation occurs on an intermediate in these preparations, after amixture is subjected to separation procedures, the chiral center isidentified with “abs” near that center, with the understanding that theseparated enantiomers may not be enantiomerically pure and the specificorientation of that bond is not drawn because the enantiomer was notconfirmed. Typically, the enriched enantiomer at each chiral centeris >90% of the isolated material. Efforts are also undertaken to enrichthe enantiomeric purity at a center to be >98% of the mixture and even>99%.

The optical rotation of an enantiomer can be measured using apolarimeter. According to its observed rotation data (or its specificrotation data), an enantiomer with a clockwise rotation was designatedas the (+)-enantiomer and an enantiomer with a counter-clockwiserotation was designated as the (−)-enantiomer. Racemic compounds areindicated either by the absence of drawn or described stereochemistry,or by the presence of (+/−) adjacent to the structure; in this lattercase, indicated stereochemistry represents the relative (rather thanabsolute) configuration of the compound's substituents.

Wherein preparative TLC or silica gel chromatography have been used, oneskilled in the art may choose any combination of solvents to purify thedesired compound.

Example (1):2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)propane-1,3-dioltrifluoroacetate salt

Step 1: Synthesis of 4-(benzylamino)-5,6-dimethyl-3-nitropyridin-2-yltrifluoromethanesulfonate

To a round bottom flask under nitrogen was added5,6-dimethyl-3-nitropyridine-2,4-diol (7.56 g, 41.05 mmol) indichloromethane (300 ml). To this was added triethylamine (12.5 g, 123mmol, 17.2 ml) and the reaction cooled to 0° C. Triflic anhydride (23.2g, 82.1 mmol, 13.8 ml) was added dropwise over 8 min. The reaction wasstirred at 0° C. for 1.5 hrs. To this was added benzyl amine (4.84 g,45.2 mmol, 4.93 ml) and the reaction warmed to RT and stirred for 3 hrs.The reaction mixture was washed with water (2×100 ml) and brine (1×100ml). The organics were dried over anhydrous sodium sulfate, filtered andconcentrated. The residue was purified by silica gel chromatography(Heptane:Ethyl acetate 0-50% gradient) to provide the title compound.Yield: 12.4 g, 30.5 mmol, 74.3%. LCMS m/z 406.2 [M+H]⁺. ¹H NMR (400 MHz,CDCl₃) δ 7.34-7.44 (m, 3H), 7.28-7.31 (m, 2H), 5.24 (br. s., 1H), 4.31(d, J=5.07 Hz, 2H), 2.47 (s, 3H), 2.12-2.20 (m, 3H).

Step 2: Synthesis ofN4-benzyl-5,6-dimethyl-3-nitro-N2-(2,4,4-trimethylpentan-2-yl)pyridine-2,4-diamine

A round bottom flask was charged with added4-(benzylamino)-5,6-dimethyl-3-nitropyridin-2-yltrifluoromethanesulfonate (12.4 g, 30.5 mmol) and toluene (100 ml).Triethylamine (4.63 g, 45.7 mmol, 6.38 ml) was added followed bytert-Octylamine (5.91 g, 45.7 mmol, 7.34 ml). Reaction was heated at 75°C. for 16 hrs. tert-Octylamine (5.91 g, 45.7 mmol, 7.34 ml) was addedand the reaction was heated at 75° C. for 48 hrs. The solution wasconcentrated on Celite®, and purified by silica gel chromatography(Heptane:Ethyl Acetate 0-50% gradient.) to provide the title compound asa red oil. Yield: 8.93 g, 23.2 mmol, 76.2%. LCMS m/z 386.4 [M+H]⁺. ¹HNMR (400 MHz, CDCl₃) δ 8.76 (s, 1H), 8.38 (br. s., 1H), 7.32-7.39 (m,2H), 7.27-7.31 (m, J=3.12, 3.12 Hz, 3H), 4.46 (d, J=4.29 Hz, 2H), 2.33(s, 3H), 2.14 (s, 3H), 1.99 (s, 2H), 1.56 (s, 6H), 0.98 (s, 9H).

Step 3: Synthesis of5,6-dimethyl-N2-(2,4,4-trimethylpentan-2-yl)pyridine-2,3,4-triamine

To a round bottom flask withN4-benzyl-5,6-dimethyl-3-nitro-N2-(2,4,4-trimethylpentan-2-yl)pyridine-2,4-diamine(8.90 g, 23.2 mmol) and ethanol (150 ml) was added ammonium formate(14.6 g, 231 mmol). Palladium on carbon (200 mg, 30% Pd) was added andthe reaction was stirred at 55° C. for 2 hrs. The reaction mixture wasthen cooled to RT and filtered through Celite® and the filtrateconcentrated. The residue was stirred in ethyl acetate for 1 hr thensolids removed by filtration through Celite®. The filtrate wasconcentrated to provide the title compound as an orange gum. Yield: 5.6g, 21.2 mmol, 91.5%. LCMS m/z 265.3 [M+H]⁺. ¹H NMR (600 MHz, CDCl₃) δ8.62 (s, 1H), 2.40 (s, 3H), 1.98 (s, 3H), 1.66 (s, 2H), 1.29 (s, 6H),1.05 (s, 9H).

Step 4: Synthesis of2-(ethoxymethyl)-6,7-dimethyl-N-(2,4,4-trimethylpentan-2-yl)-1H-imidazo[4,5-c]pyridin-4-amine

A solution of5,6-dimethyl-N2-(2,4,4-trimethylpentan-2-yl)pyridine-2,3,4-triamine(5.60 g, 21.2 mmol) and dichloromethane (100 ml) was cooled to 0° C. Tothis was added 2-ethoxyacetyl chloride (2.73 g, 22.2 mmol, 2.44 ml)followed by triethylamine (3.21 g, 31.8 mmol, 4.43 ml). The reaction wasstirred at 0° C. for 1.5 hrs. The reaction was diluted withdichloromethane (100 ml) and the organics washed with water (2×50 ml).The organics were dried over anhydrous sodium sulfate, filtered andconcentrated. The residue was diluted with ethanol (100 ml) and sodiumhydroxide (5.08 g, 127 mmol, 8.47 ml, 15N) was added. The reactionmixture was heated at 75° C. for 16 hrs. The reaction was cooled to RTand diluted with ethyl acetate and washed with water (2×). The combinedaqueous was washed with ethyl acetate. The combined organics were driedover anhydrous sodium sulfate, filtered and concentrated. The residuewas purified by silica gel chromatography (Heptane:Ethyl Acetate, 0-100%gradient) to provide the title compound. Yield: 1.40 g, 4.21 mmol,19.8%. LCMS m/z 333.1 [M+H]⁺. ¹H NMR (400 MHz, CDCl₃) δ 9.10 (br. s.,1H), 4.96 (br. s., 1H), 4.73 (s, 2H), 3.65 (q, J=7.02 Hz, 2H), 2.42 (s,3H), 2.25 (s, 3H), 2.07 (s, 2H), 1.59 (s, 6H), 1.29 (t, J=7.02 Hz, 3H),0.99 (s, 9H).

Step 5: Synthesis of1-((2,2-dimethyl-1,3-dioxan-5-yl)methyl)-2-(ethoxymethyl)-6,7-dimethyl-N-(2,4,4-trimethylpentan-2-yl)-1H-imidazo[4,5-c]pyridin-4-amine

To a solution of2-(ethoxymethyl)-6,7-dimethyl-N-(2,4,4-trimethylpentan-2-yl)-1H-imidazo[4,5-c]pyridin-4-amine(125 mg, 0.376 mmol), (2,2-dimethyl-1,3-dioxan-5-yl)methanol (68.7 mg,0.470 mmol) in toluene (2 ml) was addedcyanomethylenetributylphosphorane (136 mg, 0.564 mmol, 0.564 ml, 1M intoluene) and the reaction was heated at 90° C. for 1.5 hrs then cooledto RT and stirred for 16 hrs. Cyanomethylenetributylphosphorane (136 mg,0.564 mmol, 0.564 ml, 1M in toluene) was added and the reaction stirredat 90° C. for 1.5 hrs. The reaction mixture was absorbed on silica geland purified by silica gel chromatography (Heptane:Ethyl acetate 0-100%gradient.) to provide the title compound. Yield: 72 mg, 0.156 mmol, 42%.LCMS m/z 461.3 [M+H]⁺. ¹H NMR (400 MHz, CDCl₃) δ 5.13 (s, 1H), 4.79 (s,2H), 4.62 (d, J=7.81 Hz, 2H), 4.02 (dd, J=2.93, 12.29 Hz, 2H), 3.60 (q,J=7.02 Hz, 2H), 3.51 (d, J=10.93 Hz, 2H), 2.43 (s, 3H), 2.38 (s, 3H),2.06 (s, 2H), 1.90-1.98 (m, 1H), 1.58 (s, 6H), 1.47 (s, 6H), 1.24 (t,J=7.02 Hz, 3H), 0.99 (s, 9H).

Step 6: Synthesis of Example 1:2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)propane-1,3-dioltrifluoroacetate salt

A solution of1-((2,2-dimethyl-1,3-dioxan-5-yl)methyl)-2-(ethoxymethyl)-6,7-dimethyl-N-(2,4,4-trimethylpentan-2-yl)-1H-imidazo[4,5-c]pyridin-4-amine(72 mg, 0.16 mmol) in a 4:1 mixture of dichloromethane:trifluoroaceticacid (5 ml) was stirred at RT for 30 min. Methanol (10 ml) was added andthe reaction stirred at RT for 1.5 hrs. The reaction was concentratedand the residue was dissolved in dimethyl sulfoxide (1 ml) and purifiedby reversed phase HPLC. (Column: Waters Sunfire C18 19×100, 5u; Mobilephase A: 0.05% TFA in water (v/v); Mobile phase B: 0.05% TFA inacetonitrile (v/v); Gradient: HOLD at 95.0% H₂/5.0% Acetonitrile for 1.0min, 95.0% H₂O/5.0% Acetonitrile linear to 0% H₂O/100% Acetonitrile in9.0 min, HOLD at 0% H₂O/100% Acetonitrile to 10.0 min. Flow: 25mL/min.). Yield: 18.1 mg, 0.043 mmol, 27% HPLC Retention Time: 1.17 min(Column: Waters Atlantis® dc18 4.6×50, 5u; Mobile phase A: 0.05% TFA inwater (v/v); Mobile phase B: 0.05% TFA in acetonitrile (v/v); Gradient:95.0% H₂O/5.0% Acetonitrile linear to 5% H₂O/95% Acetonitrile in 4.0min, HOLD at 5% H₂O/95% Acetonitrile to 5.0 min. Flow: 2 mL/min.); HPLCm/z 309.4 [M+H]⁺.

Example (2):2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol

Step 1: Synthesis of 5,6-dimethyl-3-nitropyridine-2,4-diol

To 5,6-dimethylpyridine-2,4-diol (40.0 g, 287 mmol) (Org. Lett., 2003, 5(25), pp 4779-4782) at 18° C. was added concentrated sulfuric acid (174ml). The reaction was cooled to 0° C. at which point nitric acid(68-70%, 45.8 ml) was added over 1.5 hrs, maintaining internaltemperature below 10° C. After addition was complete the reaction wasstirred at 10° C. for 30 min. This reaction was combined with a secondreaction from 40 g 5,6-dimethylpyridine-2,4-diol. The combined reactionmixture was poured into ice-water (2 L). The yellow precipitate wascollected by filtration and washed with water (5×200 ml) and MTBE (5×100ml). The collected solids were dried under vacuum to provide the titlecompound as a yellow solid. Combined Yield: 50 g, 271.7 mmol, 47% yieldbased on 80 g starting pyridine. ¹H NMR (400 MHz, DMSO-d₆) 12.34 (br s,1H), 11.90 (br s, 1H), 2.21 (s, 3H), 1.90 (s, 3H).

Step 2: Synthesis ofN2,N2-bis(4-methoxybenzyl)-5,6-dimethyl-3-nitro-N4-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)pyridine-2,4-diamine

To a solution of 5,6-dimethyl-3-nitropyridine-2,4-diol (70.0 g, 380.1mmol) in dichloroethane (1.4 L) cooled to 0° C. was added triethylamine(80.8 g, 798 mmol). Trifluoromethanesulfonic anhydride (220 g, 779 mmol)was added over 30 min at 0° C. The reaction was stirred at 0° C. for 1.5hr. Triethylamine (42.3 g, 418 mmol) was added followed by portionwiseaddition of (2,2,5-trimethyl-1,3-dioxan-5-yl)methanamine (72.6 g, 456mmol) (Prepared from Organic & Biomolecular Chemistry, 14(2), 483-494;2016). The reaction was stirred at 0° C. for 20 min then stirred at 15°C. for 18 hrs. The reaction was cooled to 0° C. at which pointtriethylamine (115 g, 1.14 mol) was added followed byBis(4-methoxybenzyl)amine (127 g, 494 mmol). The reaction was thenstirred at 50° C. for 12 hrs. The solvent was removed and the residuewas purified via silica gel column chromatography (Petroleum Ether:EthylAcetate gradient 0-10%). Product fractions were collected and evaporatedto 10% volume. Solids were collected via filtration and filter cakewashed with petroleum ether (3×50 ml). The filtrate was concentrated andpurified via silica gel column chromatography (Petroleum Ether:EthylAcetate gradient 0-10%). Product fractions were collected and evaporatedto 10% volume. Solids were collected via filtration and the filter cakewashed with petroleum ether (3×20 ml), providing the title compound as ayellow solid. Yield: 98 g, 173.6 mmol, 45.7%. LCMS m/z 564.9 [M+H]⁺. ¹HNMR (400 MHz, CDCl₃) δ 7.05 (d, J=8.78 Hz, 4H), 6.80 (d, J=8.78 Hz, 4H),6.47 (t, J=6.15 Hz, 1H), 4.34 (s, 4H), 3.79 (s, 6H), 3.54-3.67 (m, 4H),3.42 (d, J=6.02 Hz, 2H), 2.35 (s, 3H), 2.21 (s, 3H), 1.43 (s, 3H), 1.41(s, 3H), 0.83 (s, 3H).

Step 3: Synthesis ofN2,N2-bis(4-methoxybenzyl)-5,6-dimethyl-N4-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)pyridine-2,3,4-triamine

To a solution ofN2,N2-bis(4-methoxybenzyl)-5,6-dimethyl-3-nitro-N4-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)pyridine-2,4-diamine(57.0 g, 100.9 mmol) in methanol (673 ml) was added ammonium formate(63.7 g, 1.01 mol) and then Zinc Dust (66.0 g, 1.01 mol). The reactionwas stirred for 10 min at 15° C. The reaction mixture was filteredthrough Celite® and the filtrate concentrated. The residue was dissolvedin ethyl acetate and water slowly added to form a white precipitate. Theaqueous layer was extracted with ethyl acetate. The combined organicswere washed with brine, dried over anhydrous sodium sulfate, filteredand concentrated to provide the title compound as a brown oil. Usedwithout further purification Yield: 50.0 g, 96.4 mmol, 95.5%.

LCMS m/z 535.0 [M+H]⁺.

Step 4: Synthesis ofN-(2-(bis(4-methoxybenzyl)amino)-5,6-dimethyl-4-(((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)amino)pyridin-3-yl)-2-ethoxyacetamide

To a solution ofN2,N2-bis(4-methoxybenzyl)-5,6-dimethyl-N4-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)pyridine-2,3,4-triamine(100 g, 192.8 mmol) in dichloromethane (1 L) was added triethylamine(97.5 g, 964 mmol, 134 ml). The reaction was cooled to 0° C. at whichpoint 2-ethoxyacetyl chloride (37.8 g, 308 mmol) was added dropwise. Theice bath was removed and the reaction stirred at 25° C. for 16 hrs. Thesolvent was removed, and the product used without further purification.Yield: 150 g, 192.8 mmol, assumed quantitative.

Step 5: Synthesis of2-(ethoxymethyl)-N,N-bis(4-methoxybenzyl)-6,7-dimethyl-1-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)-1H-imidazo[4,5-c]pyridin-4-amine

To a solution of crudeN-(2-(bis(4-methoxybenzyl)amino)-5,6-dimethyl-4-(((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)amino)pyridin-3-yl)-2-ethoxyacetamidefrom the above reaction in ethanol (3.45 L cooled to 0° C.) was addedsodium hydroxide (64.4 ml, 15N aqueous). After addition the reaction washeated to reflux for 16 hrs. The reaction was cooled to 15° C. at whichpoint a white precipitate formed. The solids were filtered and thefilter cake washed with water and MTBE. The white solids were dissolvedin ethyl acetate (1 L) and the organic layer was washed with brine,dried over anhydrous sodium sulfate, filtered and concentrated. Theresidue was purified by silica gel chromatography (EthylAcetate:Petroleum Ether gradient 0-45%.) to provide product. Thismaterial was combined with an additional 7.2 g of product from adifferent batch and stirred for 20 min in a 2:1 solution of petroleumether:MTBE. The solids were filtered and dried on vacuum to provide thetitle compound as a white solid. Combined yield: 94.52 g, 157 mmol, 76%yield. LCMS m/z 603.0 [M+H]⁺. ¹H NMR (600 MHz, DMSO-d₆) δ 7.16 (d,J=8.22 Hz, 4H), 6.83 (d, J=8.80 Hz, 4H), 4.82-5.27 (m, 6H), 4.36-4.80(m, 4H), 3.70 (s, 6H), 3.51-3.68 (m, 2H), 3.40-3.46 (m, 2H), 2.41 (s,3H), 2.34 (s, 3H), 1.39 (s, 3H), 1.34 (s, 3H), 1.07 (t, J=7.04 Hz, 3H),0.56 (s, 3H).

Step 6: Synthesis of Example (2):2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol

To a solution of2-(ethoxymethyl)-N,N-bis(4-methoxybenzyl)-6,7-dimethyl-1-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)-1H-imidazo[4,5-c]pyridin-4-amine(477 mg, 0.791 mmol) and dichloromethane (3 ml) in a vial was addedconcentrated hydrochloric acid (7.91 mmol, 0.66 ml) dropwise. The vialwas capped and the reaction was stirred at RT for 2 hrs then heated at50° C. for 1 hr. Additional concentrated hydrochloric acid (4.8 mmol,0.40 ml) was added and the reaction continued heating at 50° C. for 30min. The reaction was then cooled to RT and stirred for 16 hrs. Water (2ml) was added and the aqueous was washed with dichloromethane (2×3 ml).The aqueous was brought to pH 9 with solid sodium carbonate. Thereaction was then heated to reflux and cooled to 4° C. The solids werefiltered and washed with water (5 ml) and ether (5 ml) and dried undervacuum to provide the title compound as a white solid. Yield. 194 mg,0.602 mmol, 76.0%. LCMS m/z 323.3 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ5.76 (s, 2H), 4.91-5.09 (m, 1H), 4.69-4.90 (m, 2H), 4.27-4.59 (m, 3H),3.39-3.56 (m, 2H), 3.29-3.37 (m, 1H, assumed, partially obscured byH₂O), 3.14-3.29 (m, 2H), 2.95-3.13 (m, 1H), 2.41 (s, 3H), 2.29 (s, 3H),1.06-1.14 (m, 3H), 0.48 (s, 3H).

Example (3):2-((4-amino-2-(ethoxymethyl)-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol

Prepared in a manner similar to2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diolstarting from quinoline-2,4-diol. 18.5 mg prepared. LCMS m/z 345.4[M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) b ppm 0.58 (s, 3H) 1.15 (t, J=7.02 Hz,3H) 3.07-3.77 (m, 7H) 4.58-5.26 (m, 5H) 7.52 (t, J=8.0 Hz, 1H) 7.71 (t,J=8.0 Hz, 1H) 7.79 (d, J=8.0 Hz, 1H) 8.74 (d, J=8.0 Hz, 1H) 9.19 (br s,1H)

Example (4):2-((4-amino-2-(ethoxymethyl)-7,8-dihydrocyclopenta[b]imidazo[4,5-d]pyridin-1(6H)-yl)methyl)-2-methylpropane-1,3-diol

Prepared in a manner similar to2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diolstarting from 6,7-dihydro-5H-cyclopenta[b]pyridine-2,4-diol, 60 mgprepared. LCMS Retention Time: 0.715 min (Column: ACQUITY UPLC BEH C1850*2.1 mm, 1.7 um; Mobile phase A: 0.05% NH₄OH in water (v/v); Mobilephase B: Acetonitrile; Gradient: HOLD at 100% H₂O for 0.10 min, 100% H₂Oto 0% H₂O/100% Acetonitrile in 0.90 min, HOLD at 0% H₂O/100%Acetonitrile for 0.2 min. Flow: 1.0 mL/min.). LCMS m/z 335.3 [M+H]⁺. ¹HNMR (CDCl₃, 400 MHz), characteristic peaks: δ 5.1-5.3 (m, 2H), 4.8-5.0(m, 2H), 4.1-4.8 (m, 3H), 3.5-3.8 (m, 6H), 3.1-3.3 (m, 2H), 2.9-3.0 (m,2H), 2.1-2.2 (m, 2H), 1.26 (t, 3H, J=7.0 Hz), 0.65 (s, 3H).

Example (5):2-((4-amino-2-(ethoxymethyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diolformate salt

Prepared in a manner similar to2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diolstarting from 6,7-dihydro-5H-cyclopenta[b]pyridine-2,4-diol, 34 mgprepared. LCMS Retention Time: 0.621 min (Column: ACQUITY UPLC BEH C1850*2.1 mm, 1.7 um; Mobile phase A: 0.05% NH₄OH in water (v/v); Mobilephase B: Acetonitrile; Gradient: 95% H₂O/5% Acetonitrile to 0% H₂/100%Acetonitrile over 1 min, HOLD at 0% H₂O/100% Acetonitrile for 0.2 min.Flow: 1.0 mL/min.). LCMS m/z 349.2 [M+H]⁺. ¹H NMR (Methanol-d4, 400MHz), characteristic peaks: δ 8.4-8.6 (m, 1H), 4.63 (s, 2H), 3.59-3.68(m, 2H), 3.1-3.6 (m, 5H, assumed, partially obscured by solvent),2.8-3.0 (m, 3H), 1.72-2.06 (m, 4H), 1.22 (t, 3H, J=7.0 Hz), 0.60 (s,3H).

Examples (6) and (7):(R)-3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-oland(S)-3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-ol

Step 1: Synthesis of (2,2,5-trimethyl-1,3-dioxan-5-yl)methanol

To a stirred solution of 2-(hydroxymethyl)-2-methylpropane-1,3-diol(50.0 g, 416.2 mmol) and 2,2-dimethoxypropane (65.0 g, 624.0 mmol) wasadded acetone (46 ml) and p-toluenesulfonic acid (3.96 g, 20.8 mmol).The reaction mixture was heated at 30° C. for 12 hrs. The reaction wascooled to RT and a solution of sodium bicarbonate (12 g) in water (400ml) was added. The aqueous was extracted with ethyl acetate and theorganic layer was washed with brine, dried over anhydrous sodiumsulfate, filtered and concentrated to provide the title compound as awhite solid. Yield: 52.9 g, 330.2 mmol, 79.3%. ¹H NMR (400 MHz, DMSO-d₆)δ 4.58 (t, J=5.40 Hz, 1H), 3.53-3.57 (m, 2H), 3.41-3.45 (m, 2H),3.33-3.36 (m, 2H, partial obscured by solvent), 1.32 (s, 3H), 1.26 (s,3H), 0.74 (s, 3H).

Step 2: Synthesis of 5-(methoxymethyl)-2,2,5-trimethyl-1,3-dioxane

To a suspension of sodium hydride (33.8 g, 844 mmol, 60% dispersion inmineral oil) in toluene (1.35 L) cooled to 0° C. was added(2,2,5-trimethyl-1,3-dioxan-5-yl)methanol (67.6 g, 421.9 mmol) and thereaction was stirred at 0° C. for 10 min and then stirred to 40° C. for18 hrs. The reaction was cooled to 0° C., methyl iodide (135 g, 951.1mmol) was added and the reaction stirred at 15° C. for 60 h. Thereaction mixture was diluted with water (300 ml) and the aqueousextracted with petroleum ether (3×100 ml). The combined organic layerwas concentrated to provide the title compound as a yellow oil. Yield:60.0 g, 344.4 mmol, 81.6%. ¹H NMR (400 MHz, DMSO-d₆) δ 3.51-3.57 (m,2H), 3.43-3.49 (m, 2H), 3.28 (s, 2H), 3.25 (s, 3H), 1.33 (s, 3H), 1.27(s, 3H), 0.79 (s, 3H).

Step 3: Synthesis of 2-(methoxymethyl)-2-methylpropane-1,3-diol

To a suspension of 5-(methoxymethyl)-2,2,5-trimethyl-1,3-dioxane (55.0g, 315.7 mmol) in methanol (316 ml) cooled to 0° C. was addedhydrochloric acid (31.6 ml, 3M aqueous). The reaction mixture wasstirred at 25° C. for 30 min. The reaction mixture was concentrated andto the residue was added water (100 ml). The aqueous was extracted withpetroleum ether (3×200 ml) and then lyophilized to provide the titlecompound as a yellow oil. Yield: 32.2 g, 239.8 mmol, 75.9%. ¹H NMR (400MHz, CDCl₃) δ 3.65-3.69 (m, 2H), 3.62 (s, 2H), 3.54-3.58 (m, 2H), 3.37(s, 2H), 3.34 (s, 3H), 0.81 (s, 3H).

Step 4: Synthesis of3-((tert-butyldimethylsilyl)oxy)-2-(methoxymethyl)-2-methylpropan-1-ol

To a solution of 2-(methoxymethyl)-2-methylpropane-1,3-diol (515 mg,3.84 mmol) in tetrahydrofuran (25 ml) cooled to 0° C., was added sodiumhydride (144 mg, 3.61 mmol, 60% dispersion in mineral oil). The reactionwas stirred at 0° C. for 15 min. tert-Butyldimethylsilyl chloride (579mg, 3.84 mmol) in tetrahydrofuran (5 ml) was added dropwise. Thereaction mixture was stirred at 0° C. for 30 min and at RT for 3 hrs.The reaction was then diluted with methanol (1 ml), saturated aqueoussodium bicarbonate (10 ml) and water (15 ml). The aqueous was extractedwith dichloromethane (2×40 ml). The combined organics were dried overanhydrous sodium sulfate, filtered and concentrated. The residue waspurified by silica gel chromatography (Heptane:Ethyl Acetate, gradient0-60%) to provide the title compound, Yield: 446 mg, 1.80 mmol, 46.8%.¹H NMR (400 MHz, CDCl₃) δ 3.55-3.64 (m, 4H), 3.37 (d, J=3.90 Hz, 2H),3.35 (s, 3H), 0.90-0.91 (s, 9H), 0.82 (s, 3H), 0.05-0.08 (s, 6H).

Step 5: Synthesis of1-(3-((tert-butyldimethylsilyl)oxy)-2-(methoxymethyl)-2-methylpropyl)-2-(ethoxymethyl)-6,7-dimethyl-N-(2,4,4-trimethylpentan-2-yl)-1H-imidazo[4,5-c]pyridin-4-amine

To a solution of2-(ethoxymethyl)-6,7-dimethyl-N-(2,4,4-trimethylpentan-2-yl)-1H-imidazo[4,5-c]pyridin-4-amine(69 mg, 0.21 mmol) and3-((tert-butyldimethylsilyl)oxy)-2-(methoxymethyl)-2-methylpropan-1-ol(111 mg, 0.42 mmol) in toluene (1 ml) was addedcyanomethylenetributylphosphorane (125 mg, 0.519 mmol, 0.519 ml, 1.0M inacetonitrile). The reaction was stirred at 100° C. for 16 hrs. Thereaction mixture was absorbed on silica gel and purified by silica gelchromatography (Heptane:Ethylacetate, gradient 0-60%) to provide thetitle compound. Yield: 69 mg, 0.12 mmol, 59%. LCMS m/z 563.7 [M+H]⁺. ¹HNMR (400 MHz, MeOD) δ 4.38-4.78 (m, 4H), 3.44-3.65 (m, 4H), 3.02-3.22(m, 2H), 2.47 (s, 3H), 2.41 (s, 3H), 2.08 (br. s., 2H), 1.56 (s, 6H),1.25-1.35 (m, 4H), 1.19 (t, J=6.83 Hz, 3H), 0.94 (s, 9H), 0.92 (s, 9H),0.69 (s, 3H), 0.07 (s, 6H).

Step 6: Synthesis of Example (6) and (7)(R)-3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-oland(S)-3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-

To a solution of1-(3-((tert-butyldimethylsilyl)oxy)-2-(methoxymethyl)-2-methylpropyl)-2-(ethoxymethyl)-6,7-dimethyl-N-(2,4,4-trimethylpentan-2-yl)-1H-imidazo[4,5-c]pyridin-4-amine(69 mg, 0.12 mmol) in hexafluoroisopropanol (5.0 ml) was addedmethanesulfonic acid (70.7 mg, 0.735 mmol, 0.048 ml). The reaction wasstirred for 5 hrs at RT. The reaction mixture was diluted with saturatedaqueous sodium bicarbonate and washed with ethyl acetate (1×) anddichloromethane (1×). Organics were combined, dried over anhydroussodium sulfate, filtered and concentrated. The residue was purified bysilica gel chromatography (Dichloromethane:Methanol, 0-30% gradient).The product was collected, concentrated and filtered through a nylondisk. Solvent was removed to provide 41 mg of racemate. The racemate wasthen dissolved in 1 ml ethanol and purified by supercritical fluidchromatography. (Column: Phenomenex Lux Cellulose 4 5 um 21×250 mm);Mobile phase A: Methanol w/0.2% Ammonium Hydroxide (v/v); Mobile phaseB: CO₂ (v/v); Gradient: 70.0% CO₂/30.0% Methanol w/0.2% AmmoniumHydroxide Isocratic over 5 min. Flow: 75 mL/min. Back Pressure: 120Bar), to afford Ent 1 as Example (6): 13.8 mg, 0.041 mmol, 33.6%, 99%ee, and Ent 2 as Example (7): 13.1 mg, 0.039 mmol, 31.9%, 99% ee. SFCretention time Ent 1 as Example (6): 3.04 min, m/z 337.5 [M+H]⁺, SFCretention time Ent 2 as Example (7): 4.15 min, m/z 337.5 [M+H]⁺ (Column:Phenomenex Lux Cellulose 4 5 um 4.6×100 mm; Mobile phase A: Methanolw/0.2% Ammonium Hydroxide (v/v); Mobile phase B: CO₂ (v/v); Gradient:60.0% CO₂/40.0% Methanol w/0.2% Ammonium Hydroxide Isocratic over 5 min.Flow: 1.5 mL/min. Back Pressure: 120 Bar). The specific stereochemistryof Example (6) and Example (7) is not assigned. but each enantiomer is99% ee as provided above.

Example (8):2-((4-amino-6,7-dimethyl-2-(2,2,2-trifluoroethyl)-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-dioltrifluoroacetate salt

Step 1: Synthesis ofN-(2-(bis(4-methoxybenzyl)amino)-5,6-dimethyl-4-(((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)amino)pyridin-3-yl)-3,3,3-trifluoropropanamide

To a solution of 3,3,3-trifluoropropionic acid (15.1 mg, 0.118 mmol,0.0104 ml) was addedN2,N2-bis(4-methoxybenzyl)-5,6-dimethyl-N4-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)pyridine-2,3,4-triamine(60 mg, 0.11 mmol) and triethylamine (22.7 mg, 0.224 mmol, 0.031 ml).Propylphosphonic acid anhydride (143 mg, 0.224 mmol, 0.101 ml, 50% inethyl acetate) was added and the reaction stirred at RT for 1 hr. Waterwas added and the aqueous washed twice with ethyl acetate. The organicswere combined and dried over anhydrous sodium sulfate, filtered andconcentrated. The crude compound was used in the subsequent step withoutfurther purification or analysis, Yield: 60 mg, 0.093 mmol, 83%.

Step 2: Synthesis ofN,N-bis(4-methoxybenzyl)-6,7-dimethyl-2-(2,2,2-trifluoroethyl)-1-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)-1H-imidazo[4,5-c]pyridin-4-amine

Toa solution ofN-(2-(bis(4-methoxybenzyl)amino)-5,6-dimethyl-4-(((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)amino)pyridin-3-yl)-3,3,3-trifluoropropanamide(40 mg, 0.062 mmol) was added carbon tetrachloride (1 ml). To this wasadded triethylamine (18.8 mg, 0.186 mmol, 0.026 ml) andtriphenylphosphine (48.8 mg, 0.186 mmol). The reaction was heated at 80°C. for 16 hrs. The reaction mixture was then cooled to RT and filteredthrough Celite® and filter cake washed with ethyl acetate. The filtratewas concentrated, and the residue was purified by silica gelchromatography (Heptane:Ethyl Acetate, gradient 0-30%) to provide titlecompound. Yield: 10 mg, 0.016 mmol, 26%. ¹H NMR (400 MHz, CDCl₃) δ7.27-7.30 (m, 4H, assumed, partially obscured by residual CHCl₃), 6.82(d, J=8.59 Hz, 4H), 5.25-5.37 (m, 2H), 4.83-5.01 (m, 3H), 4.33-4.58 (m,2H), 3.79 (s, 6H), 3.76-3.84 (m, 1H, assumed, partially obscured by peakat 3.79 ppm), 3.60-3.74 (m, 2H), 3.48-3.56 (m, 1H), 3.19-3.27 (m, 1H),2.45 (s, 3H), 2.44 (s, 3H), 1.50 (s, 3H), 1.48 (s, 3H), 0.63 (s, 3H).LCMS m/z 627.5 [M+H]⁺.

Step 3: Synthesis of Example (8):2-((4-amino-6,7-dimethyl-2-(2,2,2-trifluoroethyl)-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-dioltrifluoroacetate salt

To a solution ofN,N-bis(4-methoxybenzyl)-6,7-dimethyl-2-(2,2,2-trifluoroethyl)-1-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)-1H-imidazo[4,5-c]pyridin-4-amine(10 mg, 0.016 mmol) in hexafluoroisopropanol (0.5 ml) was addedmethanesulfonic acid (2 drops). The reaction was stirred at RT for 3hrs. The reaction was then concentrated and the residue was diluted with1 ml dimethyl sulfoxide and purified via reversed phase HPLC (Column:Waters Sunfire C18 19×100, 5 u; Mobile phase A: 0.05% TFA in water(v/v); Mobile phase B: 0.05% TFA in acetonitrile (v/v); Gradient: 95.0%H₂O/5.0% Acetonitrile linear to 70% H₂O/30% Acetonitrile in 8.5 min to0% H₂O/100% MeCN to 9.0 min, HOLD at 0% H₂O/100% Acetonitrile from 9.0to 10.0 min. Flow: 25 mL/min.) to provide title compound, yield: 5.5 mg,0.012, 75%; HPLC Retention Time: 1.31 min. (Column: Waters Atlantis®dc18 4.6×50, 5 u; Mobile phase A: 0.05% TFA in water (v/v); Mobile phaseB: 0.05% TFA in acetonitrile (v/v); 95.0% H₂/5.0% Acetonitrile linear to5% H₂O/95% Acetonitrile in 4.0 min, HOLD at 5% H₂O/95% Acetonitrile to5.0 min. Flow: 2 mL/min.). HPLC m/z 347.5 [M+H]⁺.

Example (9):2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-ethylpropane-1,3-dioltrifluoroacetate salt

Prepared in a manner similar to Example (2), by utilizing(5-ethyl-2,2-dimethyl-1,3-dioxan-5-yl)methanol (Polymer Chemistry, 8(3),592-604; 2017), in step 5. The product was isolated by reversed-phaseHPLC (Column: Waters Sunfire C18 19×100, 5 u; Mobile phase A: 0.05% TFAin water (v/v); Mobile phase B: 0.05% TFA in acetonitrile (v/v);Gradient: 95.0% H₂O/5.0% Acetonitrile linear to 45% H₂O/55% Acetonitrilein 8.5 min to 0% H₂O/100% MeCN to 9.0 min, HOLD at 0% H₂O/100%Acetonitrile from 9.0 to 10.0 min. Flow: 25 mL/min.) to afford the titlecompound, yield: 25.2 mg, 0.056 mmol, 46.7%; HPLC Retention Time: 1.35min (Column: Waters Atlantis® dc18 4.6×50, 5 u; Mobile phase A: 0.05%TFA in water (v/v); Mobile phase B: 0.05% TFA in acetonitrile (v/v);95.0% H₂O/5.0% Acetonitrile linear to 5% H₂O/95% Acetonitrile in 4.0min, HOLD at 5% H₂O/95% Acetonitrile to 5.0 min. Flow: 2 mL/min.); HPLCm/z 337.5 [M+H]⁺.

Example (10):2-((4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol

Prepared in a manner similar to Example (1)2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diolstarting from quinoline-2,4-diol in step 1 and utilizing valeroylchloride in step 4. 37.8 mg prepared. HPLC retention time: 1.69 min(Column: Waters Atlantis® dc18 4.6×50, 5 u; Mobile phase A: 0.05% TFA inwater (v/v); Mobile phase B: 0.05% TFA in acetonitrile (v/v); Gradient:95.0% H₂O/5.0% Acetonitrile linear to 5% H₂O/95% Acetonitrile in 4.0min, HOLD at 5% H₂O/95% Acetonitrile to 5.0 min. Flow: 2 mL/min.). HPLCm/z 343.5 [M+H]⁺.

Example (11):2-((4-amino-2-butyl-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol

Prepared in a manner similar to2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diolutilizing valeroyl chloride in Example 1, step 4. 260 mg prepared. ¹HNMR (DMSO-d₆, 400 MHz) δ 5.56 (s, 2H), 4.8-5.0 (m, 2H), 4.3-4.5 (m, 1H),4.1-4.3 (m, 1H), 3.1-3.3 (m, 3H), 3.0-3.1 (m, 1H), 2.8-3.0 (m, 2H), 2.40(s, 3H), 2.28 (s, 3H), 1.6-1.7 (m, 2H), 1.3-1.4 (m, 2H), 0.91 (t, 3H,J=7.2 Hz), 0.45 (s, 3H). LCMS m/z 321.2 [M+H]⁺.

Example (12):2-((4-amino-2-pentyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol

Prepared in a manner similar to Example (1)2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diolstarting from quinoline-2,4-diol in step 1 and utilizing Hexanoylchloride in step 4. 31.2 mg prepared. HPLC retention time: 1.88 min(Column: Waters Atlantis® dc18 4.6×50, 5 u; Mobile phase A: 0.05% TFA inwater (v/v); Mobile phase B: 0.05% TFA in acetonitrile (v/v); Gradient:95.0% H₂O/5.0% Acetonitrile linear to 5% H₂O/95% Acetonitrile in 4.0min, HOLD at 5% H₂O/95% Acetonitrile to 5.0 min. Flow: 2 mL/min.). HPLCm/z 357.5 [M+H]⁺.

Example (13):2-((4-amino-2-butyl-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diolformate salt

Step 1: Synthesis of 3-nitro-5,6,7,8-tetrahydroquinoline-2,4-diol

To a 2 L flask was added sulfuric acid (275 ml). The reaction was cooledin an ice bath and 5,6,7,8-tetrahydroquinoline-2,4-diol (65 g, 390 mmol)was added portionwise over 15 minutes. The reaction stirred for anadditional 10 minutes. Nitric acid (39.6 ml, 885 mmol) was addedportionwise at a rate maintaining the internal reaction temperaturebelow 30° C. The reaction was stirred at RT for an additional 2 hours.The reaction was slowly poured into ice (2 L), the solids filtered andwashed with water. The solids were dried under vacuum at 50° C. Yield:52.2 g, 390 mmol, 63%. ¹H NMR (400 MHz, DMSO-d₆) δ ppm 12.25 (br s, 1H),11.75 (br s, 1H), 2.48 (m, 2H), 2.34 (m, 2H), 1.66 (m, 4H). LCMS m/z211.2 [M+H].

Step 2: Synthesis of 2,4-dichloro-3-nitro-5,6,7,8-tetrahydroquinoline

To a 500 ml flask was added 3-nitro-5,6,7,8-tetrahydroquinoline-2,4-diol(13.1 g, 53.0 mmol), dichloroethane (70.0 mL), phosphorous (V)oxychloride (65.1 g, 425 mmol, 40.0 ml). The reaction was heated at 80°C. for 16 hours. The reaction was then cooled to RT and evaporatedfollowed by evaporation from toluene (2×). The residue was filteredthrough a silica plug with dichloromethane and the eluent was evaporatedto provide a brown/orange waxy solid. Yield: 9.2 g, 37.2 mmol, 70%. ¹HNMR (400 MHz, CDCl₃) δ 2.92-3.02 (m, 2H), 2.77-2.85 (m, 2H), 1.85-1.95(m, 4H). GCMS m/z 246.0. Additional material was prepared using similarconditions.

Step 3: Synthesis of2-chloro-3-nitro-N-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)-5,6,7,8-tetrahydroquinolin-4-amine

To 2,4-dichloro-3-nitro-5,6,7,8-tetrahydroquinoline (10.9 g, 44.5 mmol)and dimethylacetamide (55 mL, contains 20% water). Triethylamine (9 g,89.0 mmol, 12.4 mL) and then(2,2,5-trimethyl-1,3-dioxan-5-yl)methanamine (12.7 g, 80.1 mmol) wasadded and the reaction was heated at 35° C. for 16 hours. The reactionwas cooled to 0° C., water (70 mL) added and reaction stirred for 45min. The solids were filtered and washed with water to provide titlecompound as an orange solid. Yield: 14.78 g, 39.96 mmol, 89.8%. ¹H NMR(400 MHz, CDCl₃) δ 5.57 (br. s., 1H), 3.67-3.78 (m, 4H), 3.21 (d, J=4.68Hz, 2H), 2.83 (t, J=5.66 Hz, 2H), 2.48 (t, J=5.66 Hz, 2H), 1.78-1.92 (m,4H), 1.47 (d, J=5.85 Hz, 6H), 0.87 (s, 3H). LCMS m/z 370.4 [M+H].

Step 4: Synthesis ofN2,N2-bis(4-methoxybenzyl)-3-nitro-N4-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)-5,6,7,8-tetrahydroquinoline-2,4-diamine

To2-chloro-3-nitro-N-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)-5,6,7,8-tetrahydroquinolin-4-amine(12.98 g, 35.09 mmol) was added Bis(4-methoxybenzyl)amine (27.1 g, 105mmol) and isopropanol (65 mL). The reaction was refluxed for 51 hoursthen stirred at RT for 16 hours. The reaction was the diluteddichloromethane (100 mL) and filtered through Celite®. The solids werewashed with dichloromethane (50 mL). The organics were concentrated anddiluted with ethanol (40 mL) and stirred at RT for 16 hrs. The solidswere filtered, and flask washed with ethanol (60 mL). The solids werewashed with ethanol (30 mL) and dried under vacuum to provide a yellowsolid. Yield: 14.5 g, 24.6 mmol, 70.0%. ¹H NMR (400 MHz, CDCl₃) δ7.07-7.12 (m, 4H), 6.76-6.84 (m, 4H), 6.39 (t, J=5.66 Hz, 1H), 4.30 (s,4H), 3.79 (s, 6H), 3.55-3.67 (m, 4H), 3.45 (d, J=5.85 Hz, 2H), 2.73 (t,J=6.44 Hz, 2H), 2.66 (t, J=5.85 Hz, 2H), 1.72-1.86 (m, 4H), 1.44 (s,3H), 1.41 (s, 3H), 0.82-0.88 (m, 3H). LCMS m/z 591.4 [M+H].

Step 5:N2,N2-bis(4-methoxybenzyl)-N4-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)-5,6,7,8-tetrahydroquinoline-2,3,4-triamine

ToN2,N2-bis(4-methoxybenzyl)-3-nitro-N4-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)-5,6,7,8-tetrahydroquinoline-2,4-diamine(7.2 g, 12.2 mmol) was added methanol (40.6 mL), ammonium formate (3.84g, 60.9 mmol), and zinc dust (3.99 g, 60.9 mmol). The reaction wasstirred for 25 minutes. The reaction was filtered through a pad ofCelite® and filtrate concentrated. The residue was dissolved in ethylacetate, washed with water, brine and the organic was dried overanhydrous sodium sulfate. The reaction was filtered and concentrated toprovide title compound. Yield: 6.83 g, 12.19 mmol, 100%. ¹H NMR (400MHz, CHLOROFORM-d) b ppm 7.21 (d, J=8.59 Hz, 4H), 6.81 (d, J=8.59 Hz,4H), 4.15 (s, 4H), 3.99 (s, 2H), 3.76-3.83 (m, 8H), 3.63-3.70 (m, 2H),3.21 (br s, 2H), 2.74 (br t, J=5.66 Hz, 2H), 2.55 (br t, J=5.46 Hz, 2H),1.74-1.87 (m, 4H), 1.48 (s, 3H), 1.46 (s, 3H), 0.93 (s, 3H) LCMS m/z561.5 [M+H].

Step 6: Synthesis ofN-(2-(bis(4-methoxybenzyl)amino)-4-(((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)amino)-5,6,7,8-tetrahydroquinolin-3-yl)pentanamide

ToN2,N2-bis(4-methoxybenzyl)-N4-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)-5,6,7,8-tetra-hydroquinoline-2,3,4-triamine(6.84 g, 12.19 mmol) was added dichloromethane (60.9 mL), water (30.5mL) and sodium bicarbonate (2.56 g, 30.5 mmol). Valeryl chloride (1.62g, 13.4 mmol, 1.59 mL) was added dropwise over 1 minute and then stirredfor 55 minutes. The organic layer was separated and the aqueous washedwith dichloromethane. The combined organic layers were washed withbrine, dried over anhydrous sodium sulfate, filtered and concentrated toprovide title compound which was used without further purification.Yield: 7.86 g, 12.19 mmol. LCMS m/z 645.7 [M+H].

Step 7: Synthesis of2-butyl-N,N-bis(4-methoxybenzyl)-1-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-4-amine

ToN-(2-(bis(4-methoxybenzyl)amino)-4-(((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)amino)-5,6,7,8-tetrahydroquinolin-3-yl)pentanamide(7.86 g, 12.19 mmol) was added ethanol (122 mL) and sodium hydroxide(4.88 g, 60.9 mmol, 3.22 mL, 50% wt solution in water). The reaction washeated at 100° C. for 48 hours. The reaction was cooled to RT and thesolids were filtered, washed with ethanol and dried to provide titlecompound. Yield: 6.7 g, 10.7 mmol, 87.7%. ¹H NMR (400 MHz, CHLOROFORM-d)b ppm 7.26 (d, J=8.59 Hz, 4H), 6.81 (d, J=8.59 Hz, 4H), 4.96-5.40 (m,4H), 4.39-4.72 (m, 2H), 3.79 (s, 6H), 3.41-3.69 (m, 4H), 2.63-3.13 (m,6H), 1.84 (br s, 4H), 1.69 (quin, J=7.51 Hz, 2H), 1.57 (br s, 4H), 1.46(s, 6H), 1.35 (dq, J=14.83, 7.41 Hz, 2H), 0.89 (t, J=7.41 Hz, 3H), 0.57(s, 3H). LCMS m/z 627.7 [M+H].

Step 8: Synthesis of2-((4-amino-2-butyl-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol

To2-butyl-N,N-bis(4-methoxybenzyl)-1-((2,2,5-trimethyl-1,3-dioxan-5-yl)methyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-4-amine(6.70 g, 10.69 mmol) was added toluene (32.4 mL) and concentratedhydrochloric acid (21 mL). The reaction was heated at 60° C. for 3.25hours. The aqueous layer was washed with toluene, heated to 60° C. andbrought to pH 10 with solid potassium carbonate. The reaction was thenstirred at 60° C. for 1 hour and 40 minutes followed by cooling to RT.The solids were filtered, rinsed with water and dried under vacuum at40° C. to provide Example (13). Yield: 2.44 g, 7.04 mmol, 65.9% Yield.¹H NMR (400 MHz, DMSO-d₆) b ppm 5.67 (br s, 2H), 4.78 (br s, 2H),4.07-4.45 (m, 2H), 2.71-3.30 (m, 8H), 2.66 (br s, 2H), 1.53-1.93 (m,6H), 1.35 (m, J=7.34, 7.34, 7.34, 7.34, 7.34 Hz, 2H), 0.91 (t, J=7.41Hz, 3H), 0.43 (s, 3H). LCMS m/z 347.3 [M+H].

Example (14):2-((4-amino-2-butyl-7,8-dihydrocyclopenta[b]imidazo[4,5-d]pyridin-1(6H)-yl)methyl)-2-methylpropane-1,3-diol

Prepared in a manner similar to Example 13, starting from6,7-dihydro-5H-cyclopenta[b]pyridine-2,4-diol. 336 mg prepared (42.5%Yield). Purified by HPLC (Column: Welch Xtimate 75*40 mm*3 um; Mobilephase A: 0.05% NH₄OH in water (v/v); Mobile phase B: Acetonitrile;Gradient: 80% A to 40% A/60% B in 10 min, HOLD at 0% H₂/100%Acetonitrile for 4 min. Flow: 25 mL/min.). LCMS m/z 333.1 [M+H]⁺. ¹H NMR(400 MHz, DMSO) δ 5.72 (s, 2H), 4.80 (br. s., 2H), 4.14 (s, 2H), 3.25(br. s., 3H), 2.99-3.34 (m, 3H), 2.88 (br. s., 2H), 2.62-2.78 (m, 2H),2.02 (apparent quin, J=7.34 Hz, 2H), 1.64-1.75 (m, 2H), 1.35 (apparentqd, J=7.47, 14.74 Hz, 2H), 0.91 (t, J=7.28 Hz, 3H), 0.50 (s, 3H).

Example (15):2-((4-amino-2-ethyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol

Prepared in a manner similar to2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol,starting from quinoline-2,4-diol. 301 mg prepared. Purified by HPLC(Column: Phenomenex Gemini NX-C18 150*30 mm*5 um; Mobile phase A: 0.05%NH₄OH in water (v/v); Mobile phase B: Acetonitrile; Gradient: 95% A to55% A/45% B in 7 min, HOLD at 0% H₂O/100% Acetonitrile for 2 min. Flow:30 mL/min.)¹H NMR (400 MHz, DMSO-d6) δ 8.51 (d, J=8.1 Hz, 1H), 7.58 (d,J=8.3 Hz, 1H), 7.38 (t, J=7.5 Hz, 1H), 7.18 (t, J=7.4 Hz, 1H), 6.43 (s,2H), 4.98 (br s, 2H), 4.78 (br s, 1H), 4.45 (br s, 1H), 3.19 (br s, 4H),3.02 (q, J=7.4 Hz, 2H), 1.34 (t, J=7.4 Hz, 3H), 0.55 (s, 3H). LCMS m/z315.1 [M+H]⁺.

Example (16):2-((4-amino-2-propyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol

Prepared in a manner similar to Example 13, starting fromquinoline-2,4-diol. 15.8 mg prepared. Purified by HPLC (Column: WatersXBridge C18 19×100, 5 um; Mobile phase A: 0.03% NH₄OH in water (v/v);Mobile phase B: 0.03% NH₄OH Acetonitrile; Gradient: 95% A to 50% A/50% Bin 8.5 min, HOLD at 0% H₂O/100% Acetonitrile for 1 min. Flow: 25mL/min.;). HPLC QC (Column: Waters Atlantis dC18 4.6×50, 5 um; Mobilephase A: 0.05% TFA in water (v/v); Mobile phase B: 0.05% TFAAcetonitrile; Gradient: 95% A to 5% A/95% B in 4.0 min, HOLD at 5%H₂O/95% Acetonitrile for 1 min. Flow: 2 mL/min.; Retention Time: 1.38min). LCMS m/z 329.5 [M+H]⁺.

Example (17):2-((4-amino-2-(2-methoxyethyl)-7,8-dihydrocyclopenta[b]imidazo[4,5-d]pyridin-1(6H)-yl)methyl)-2-methylpropane-1,3-diol

Example 17 was prepared in a manner similar to2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol,starting from 6,7-dihydro-5H-cyclopenta[b]pyridine-2,4-diol. 70 mgprepared. Purified by HPLC (Column: Phenomenex Gemini NX-C18 75*30 mm*3um; Mobile phase A: 0.05% NH₄OH in water (v/v); Mobile phase B:Acetonitrile; Gradient: 100% A to 70% A/30% B in 7 min, HOLD at 0%H₂O/100% Acetonitrile for 2 min. Flow: 30 mL/min.). LCMS m/z 335.1[M+H]⁺. ¹H NMR (400 MHz, DMSO) δ 5.87 (br. s., 2H), 4.81 (br. s., 2H),4.17 (br. s., 2H), 3.71 (apparent t, J=6.72 Hz, 2H), 3.30 (br. s., 3H),3.20-3.28 (m, 4H), 3.23 (s, 3H), 2.68-2.79 (m, 2H), 2.03 (quin, J=7.27Hz, 2H), 0.50 (s, 3H). 1 proton not observed (obscured).

Example (18):2-((4-amino-2-(2-methoxyethyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol

Example (18) was prepared in a manner similar to2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol,starting from 5,6,7,8-tetrahydro-quinoline-2,4-diol. 68 mg prepared.Purified by HPLC (Column: Phenomenex Gemini NX-C18 75*30 mm*3 um; Mobilephase A: 0.05% NH₄OH in water (v/v); Mobile phase B: Acetonitrile;Gradient: 100% A to 60% A/40% B in 7 min, HOLD at 0% H₂O/100%Acetonitrile for 2 min. Flow: 30 mL/min.). ¹H NMR (400 MHz, DMSO-d₆) δ5.70 (s, 2H), 4.82 (br s, 2H), 4.29 (br s, 2H), 3.69 (br s, 2H),3.30-2.72 (m, 11H), 2.66 (br s, 2H), 1.74 (br s, 4H), 0.43 (s, 3H). LCMSm/z 349.2 [M+H]⁺.

Example (19):2-((4-amino-2-(2-methoxyethyl)-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol

Example (19) was prepared in a manner similar to2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol,starting from quinoline-2,4-diol. 108 mg prepared. Purified by HPLC(Column: Phenomenex Gemini NX-C18 75*30 mm*3 um; Mobile phase A: 0.05%NH₄OH in water (v/v); Mobile phase B: Acetonitrile; Gradient: 97% A to57% A/43% B in 7 min, HOLD at 0% H₂O/100% Acetonitrile for 2 min. Flow:25 mL/min.;) LCMS m/z 345.3 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) 5=8.52 (d,J=8.3 Hz, 1H), 7.58 (dd, J=1.1, 8.3 Hz, 1H), 7.43-7.34 (m, 1H),7.23-7.14 (m, 1H), 6.45 (s, 2H), 5.00 (t, J=4.8 Hz, 2H), 4.76 (br s,1H), 4.53 (br s, 1H), 3.79 (br s, 2H), 3.43 (br d, J=5.8 Hz, 3H),3.30-3.24 (m, 4H), 3.19 (br s, 2H), 0.55 (s, 3H).

Biological Testing

The Examples described were tested for biological activity in functionalcellular assays using HEK293 cells stably overexpressing human TLR7 orTLR8. The assays tested for the ability of each Example to stimulatesecretion of interferon alpha (IFNα) in human primary blood mono nuclearcells (PBMC).

hTLR7 and hTLR8 Cell Functional Assays

To determine the ability of each Example to activate the human toll likereceptor 7 (hTLR7) or human toll like receptor 8 (hTLR8), cell-basedreporter systems were utilized. HEK293 cells stably overexpressingeither hTLR7 or hTLR8 along with a reporter gene containing an optimizedsecreted embryonic alkaline phosphatase gene (SEAP), under the controlof the IFN-b minimal promoter fused to five NF-κB and AP-1-bindingsites, were obtained from Invivogen (HEK-Blue™ hTLR7, cat #Hkb-htlr7;HEK-Blue™ hTLR8, cat #Hkb-htlr8). Stimulation of hTLR7 or hTLR8 in thesecells activates NF-κB and AP-1 and induces the production of SEAP whichcan be quantified using an alkaline phosphatase detection reagent.

Cells were maintained in Dulbecco's Modified Eagle Media (DMEM)(containing Fetal Bovine Serum (FBS) heat inactivated (10%), Glutamax (2mM), Penicillin/Streptomycin, Blasticidin (10 μg/ml), Zeocin (100 μg/ml)and Normocin (100 μg/ml)). On day one of the assay, Examples wereprepared using 11-point half-log serial dilutions from a 10 mM DMSOstock solution and 50 nl was spotted into 384-well Viewplates(PerkinElmer, cat #6007480). Positive control TLR7/8 agonist andnegative control (DMSO, no Example) were also spotted within the assayplate and were used to determine percent effect during the analysisprocess. After resuspension in DMEM assay media containing FBS heatinactivated (10%), Glutamax (2 mM) and Penicillin/Streptomycin, 10,000cells/20 μl/well were added to previously prepared plates. Plates wereincubated overnight (16-20 hrs) at 37° C. in a 5% CO₂ environment.Prewetted Microclime lids (Labcyte, LLS-0310) were used to preventevaporation. On day two of the assay, QUANTI-Blue™ detection reagent wasprepared by reconstituting QUANTI-Blue™ powder (InvivoGen, Rep-qbl) with100 ml of sterile water and allowed to equilibrate to 37° C. for 15minutes. 20 μl of QUANTI-Blue™ detection reagent was added to each welland plates were incubated at room temperature for 180 min. At the end ofthe incubation, plates were read on an Envision (Perkin Elmer) platereader capturing absorbance at 650 nm.

Using Positive (TLR7/8 agonist) and Negative (DMSO) controls, thepercent (%) effect was calculated for each Example using the followingequation:% effect=100−[100*{(Example−Positive Control)/(Negative Control−PositiveControl)}]

The % effect at each concentration of each Example was calculatedutilizing the ABase software suite (IBDS) and was relative to the amountof SEAP produced in the positive and negative control wells containedwithin each assay plate. The concentrations and % effect values for eachExample were fit using a 4-parameter logistic model in ABase and theconcentration of each Example that produced 50% response (EC₅₀) wascalculated.

INFα Assay from Peripheral Blood Mononuclear Cells (PBMC)

To determine the ability of each Example to induce the release ofinterferon alpha (IFNα) from freshly isolated peripheral bloodmononuclear cells (PBMCs), a Homogeneous Time-Resolved Fluorescence(HTRF) assay was utilized. Human whole blood was collected from healthydonors via vein puncture in accordance with Pfizer protocols (ProtocolNo. GOHW RDP-01), approved by the Shulman Institutional Review Board. 50ml of human venous blood sample from individual donors was heparinizedby addition to a conical tube containing 714 units of Heparin SodiumInjection MDV (Fresenius Kabi, cat #70041) followed by gentle inversionof the tube several times. Blood was then transferred to a flask, theconical tube was rinsed with 40 ml of PBS containing 2 mM EDTA(PBS-EDTA), and the rinse was added to the blood flask with gentlymixing. 30 ml of the diluted blood was added to 3 separate histopaquetubes (Sigma, cat #A0561), applying directly to the frit. Histopaquetubes were then spun for 15 min at 1000×g in a tabletop centrifuge.Following density separation, excess upper phase of plasma was aspiratedto within ˜5 ml above the interphase and remaining plasma along with thecloudy interphase containing PBMCs were gently decanted into a newconical tube. 15 ml of PBS-EDTA was added to the histopaque tube andswirled gently to remove remaining PBMCs that adhered to the tube walland this wash was added to the existing PBMCs in the tube. The volume ofthe tube was brought to 40 ml with PBS-EDTA and tubes were spun at 250×gfor 12 minutes at room temperature. After aspiration of the supernatant,the pellet was gently resuspended with 10 ml of PBS-EDTA and centrifugedagain at 250×g for 12 minutes. The resulting supernatant was decanted,and the pellet was resuspended in 20 ml of ACK lysing buffer(ThermoFisher, cat #A10492-01) followed by incubation at roomtemperature for 5 minutes. The volume of each tube was brought to 50 mlwith the addition of PBS-EDTA and tubes were spun at 177×g for 12minutes at room temperature. The PBMC pellet was again resuspended in 10ml of PBS without EDTA and tubes were spun for a final time at 177×g for10 minutes. The supernatant was decanted and PBMCs were resuspended inAssay Media (RPMI base media with 10% FBS heat inactivated, 2 mMGlutamax and Penicillin/Streptomycin).

Each Example was prepared for the assay using 11-point half log serialdilutions from a 2.5 mM DMSO stock solution and 400 nl was spotted into384 well Viewplate (Perkin Elmer, cat #6007480). Positive and negativecontrols (previously described) were also spotted within the assay plateand were used to determine percent effect during the analysis process.PBMCs were counted and plated at a density of 100,000 cells/100 μl/well;plates were covered with a prewet microclime lid to prevent evaporationand incubated for 24 hrs at 37° C. in a 5% CO₂ atmosphere. At the end ofthe incubation, the microclime lid was removed and the plates were spunat 1000 rpm for 5 minutes. 16 μl of conditioned media from the cellplate was transferred into a separate 384 well low-volume plate (GreinerOne, 784080). IFNα levels were quantitated using an HTRF® kit (Cisbio,cat #62HIFNAPEG) according to the manufacturer's instructions. The kitsupplies two different specific antibodies, one labelled with D2(acceptor) and the other labelled with cryptate (donor), and detectionbuffer. The antibody stocks were diluted 1:20 in detection buffer. Forone 384 well plate, 62.5 μl of D2 antibody stock and 62.5 μl of cryptateantibody stock was added to 2.375 ml of detection buffer and mixed well.4 μl of the antibody mix was added to each well containing theconditioned media obtained from the corresponding well of the Viewplate.The low-volume plates were sealed and incubated for 24 hours at roomtemperature. The HTRF signal was read with an Envision multi-label platereader (Perkin Elmer) using excitation of 330 nm and emissions of 615 nmand 665 nm. Results were calculated as (665 nm/615 nM ratio)*10,000 andraw data was converted to concentration of IFNα (pg/ml) using thecytokine standard curve. As discussed above, Positive (TLR7/8 agonist)and Negative (DMSO) controls were used to calculate the percent (%)effect for each Example using the following equation:% effect=100−[100*{(Example−Positive Control)/(Negative Control−PositiveControl)}]

The % effect at each concentration for each Example was calculatedutilizing the ABase software suite (IBDS) and was relative to the amountof IFNα produced in the positive and negative control wells containedwithin each assay plate. The concentrations and % effect values for eachExample were fit using a 4-parameter logistic model in ABase and theconcentration of Example that produced 50% response (EC₅₀) wascalculated and provided in Table 1, as the geometric mean EC₅₀ when anExample was tested more than once. A blank cell in Table 1 indicates nodata was obtained for that Example in that specific assay.

TABLE 1 Exam- hTLR7 hTLR8 IFNα ple (cell) EC₅₀ (cell) EC₅₀ (PBMC) number(μM) (μM) EC₅₀ (μM) 1 0.644 2.549 0.099 2 0.054 0.811 0.019 3 0.0290.194 0.007 4 0.054 0.400 5 0.042 0.273 6 0.049 6.398 0.022 7 0.10316.472 0.198 8 0.078 8.018 0.048 9 0.027 1.443 0.002 10 0.004 0.0920.001 11 0.013 0.160 12 0.009 0.191 13 0.009 0.104 0.0004 14 0.014 0.2470.007 15 0.075 0.613 0.012 16 0.019 0.325 0.011 17 0.156 1.064 18 0.0160.119 19 0.009 0.310

All publications and patent applications cited in the specification areherein incorporated by reference in their entirety. It will be apparentto those of ordinary skill in the art that certain changes andmodifications may be made thereto without departing from the spirit orscope of the appended claims.

The invention claimed is:
 1. A compound of Formula (I)

or a pharmaceutically acceptable salt thereof, wherein R¹ and R² areindependently C₁₋₃ alkyl; or R¹ and R² are joined to form a 5- to7-membered carbocyclic ring, wherein said carbocyclic ring may besaturated or unsaturated; R³ is

R⁴ is C₁₋₆ alkyl, or (CH₂)_(n)O(CH₂)_(m)CH₃, wherein the C₁₋₆ alkyl orany carbon of the (CH₂)_(n)O(CH₂)_(m)CH₃ group is substituted with 0 to3 halogen as valency allows; R⁵ is C₁₋₃ alkyl, or OC₁₋₃ alkyl, whereinthe C₁₋₃ alkyl is substituted by 0 to 3 F; R⁶ is H, or C₁₋₃ alkyl,wherein the C₁₋₃ alkyl is substituted with 0 to 3 F; m is 0 to 2; and nis 1 to
 3. 2. A compound of Formula (Ia)

or a pharmaceutically acceptable salt thereof, wherein R¹ and R² areindependently C₁₋₂ alkyl; or R¹ and R² are joined to form a 5- to7-membered carbocyclic ring, wherein said carbocyclic ring may besaturated or unsaturated; R³ is

R⁴ is C₃₋₅ alkyl, or (CH₂)_(n)O(CH₂)_(m)CH₃; R⁵ is C₁₋₂ alkyl; R⁶ is H;m is 1; and n is
 1. 3. The compound of claim 2, or a pharmaceuticallyacceptable salt thereof, wherein R¹ and R² are independently C₁₋₂ alkyl.4. The compound of claim 2, or a pharmaceutically acceptable saltthereof, wherein R¹ and R² are joined to form a 5- to 7-memberedcarbocyclic ring, wherein said carbocyclic ring may be saturated orunsaturated.
 5. The compound of claim 4, or a pharmaceuticallyacceptable salt thereof, wherein R¹ and R² are joined to form a 5- to7-membered carbocyclic ring, wherein said carbocyclic ring may besaturated.
 6. The compound of claim 5, or a pharmaceutically acceptablesalt thereof, wherein the carbocyclic ring is cyclopentyl.
 7. Thecompound of claim 5, or a pharmaceutically acceptable salt thereof,wherein the carbocyclic ring is cyclohexyl.
 8. The compound of claim 4,or a pharmaceutically acceptable salt thereof, wherein R¹ and R² arejoined to form a 5- to 7-membered carbocyclic ring, wherein saidcarbocyclic ring may be unsaturated; and R⁴ is C₃₋₅ alkyl.
 9. Thecompound of claim 8, or a pharmaceutically acceptable salt thereof,wherein the carbocyclic ring is phenyl.
 10. A compound of Formula (Ib)

or a pharmaceutically acceptable salt thereof, wherein R¹ and R² areindependently C₁₋₃ alkyl; or R¹ and R² are joined to form a 5- to7-membered carbocyclic ring, wherein said carbocyclic ring may besaturated or unsaturated; R³ is

R⁴ is C₁₋₆ alkyl, or (CH₂)_(n)O(CH₂)_(m)CH₃, wherein said C₁₋₆ alkyl orany carbon of the (CH₂)_(n)O(CH₂)_(m)CH₃ group is substituted with 0 to3 halogen as valency allows, wherein halogen is F; R⁵ is C₁₋₃ alkyl, orOC₁₋₃ alkyl, wherein the C₁₋₃ alkyl is substituted by 0 to 3 F; R⁶ is H,or C₁₋₃ alkyl, wherein the C₁₋₃ alkyl is substituted with 0 to 3 F; m is0 to 2; and n is 1 to
 3. 11. The compound of claim 10, or apharmaceutically acceptable salt thereof, wherein R¹ and R² areindependently C₁₋₂ alkyl; or R¹ and R² are joined to form a 5- to7-membered carbocyclic ring, wherein said carbocyclic ring may besaturated or unsaturated; R³ is

R⁵ is C₁₋₃ alkyl, or OC₁₋₃ alkyl, wherein the C₁₋₃ alkyl is substitutedby 0 to 2 F; and R⁶ is H.
 12. The compound of claim 11, or apharmaceutically acceptable salt thereof, wherein R⁵ is C₁₋₂ alkyl. 13.A compound selected from:2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol;2-((4-amino-2-(ethoxymethyl)-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;2-((4-amino-2-(ethoxymethyl)-7,8-dihydrocyclopenta[b]imidazo[4,5-d]pyridin-1(6H)-yl)methyl)-2-methylpropane-1,3-diol;2-((4-amino-2-(ethoxymethyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-ol;(R)-3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-ol;(S)-3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-ol;2-((4-amino-6,7-dimethyl-2-(2,2,2-trifluoroethyl)-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol;2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-ethylpropane-1,3-diol;2-((4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;2-((4-amino-2-butyl-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol;and2-((4-amino-2-pentyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;or a pharmaceutically acceptable salt thereof.
 14. A compound that is2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol;or a pharmaceutically acceptable salt thereof.
 15. The compound of claim14 that is2-((4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)methyl)-2-methylpropane-1,3-diol.16. A compound that is2-((4-amino-2-(ethoxymethyl)-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;or a pharmaceutically acceptable salt thereof.
 17. The compound of claim16 that is2-((4-amino-2-(ethoxymethyl)-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol.18. A compound that is2-((4-amino-2-(ethoxymethyl)-7,8-dihydrocyclopenta[b]imidazo[4,5-d]pyridin-1(6H)-yl)methyl)-2-methylpropane-1,3-diol;or2-((4-amino-2-(ethoxymethyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;or a pharmaceutically acceptable salt thereof.
 19. The compound of claim18 that is2-((4-amino-2-(ethoxymethyl)-7,8-dihydrocyclopenta[b]imidazo[4,5-d]pyridin-1(6H)-yl)methyl)-2-methylpropane-1,3-diol;or2-((4-amino-2-(ethoxymethyl)-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol.20. A compound that is3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-01;(R)-3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-ol;or(S)-3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-ol;or a pharmaceutically acceptable salt thereof.
 21. The compound of claim20 that is3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-ol;(R)-3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-ol;or(S)-3-(4-amino-2-(ethoxymethyl)-6,7-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)-2-(methoxymethyl)-2-methylpropan-1-ol.22. A compound that is2-((4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;or2-((4-amino-2-pentyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;or a pharmaceutically acceptable salt thereof.
 23. The compound of claim22 that is2-((4-amino-2-butyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;or2-((4-amino-2-pentyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol.24. A pharmaceutical composition comprising the compound of claim 1, ora pharmaceutically acceptable salt thereof, and a pharmaceuticallyacceptable carrier or excipient.
 25. A compound selected from2-((4-amino-2-butyl-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;2-((4-amino-2-butyl-7,8-dihydrocyclopenta[b]imidazo[4,5-d]pyridin-1(6H)-yl)methyl)-2-methylpropane-1,3-diol;2-((4-amino-2-ethyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;and2-((4-amino-2-propyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol;or a pharmaceutically acceptable salt thereof.
 26. A compound of claim25, or a pharmaceutically acceptable salt thereof, that is2-((4-amino-2-butyl-6,7,8,9-tetrahydro-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol.27. A compound of claim 25, or a pharmaceutically acceptable saltthereof, that is2-((4-amino-2-butyl-7,8-dihydrocyclopenta[b]imidazo[4,5-d]pyridin-1(6H)-yl)methyl)-2-methylpropane-1,3-diol.28. A compound of claim 25, or a pharmaceutically acceptable saltthereof, that is2-((4-amino-2-ethyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol.29. A compound of claim 25, or a pharmaceutically acceptable saltthereof, that is2-((4-amino-2-propyl-1H-imidazo[4,5-c]quinolin-1-yl)methyl)-2-methylpropane-1,3-diol.30. A pharmaceutical composition comprising the compound of claim 25, ora pharmaceutically acceptable salt thereof, and a pharmaceuticallyacceptable carrier or excipient.